Diarymethyl and related compounds

ABSTRACT

This invention provides compounds of formula I: 
     
       
         
         
             
             
         
       
     
     wherein R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , Ar 1 , Ar 2 , E, a, b, c and z are as defined in the specification, or a pharmaceutically acceptable salt or solvate or stereoisomer thereof. The compounds of this invention possess both β 2  adrenergic receptor agonist and muscarinic receptor antagonist activity and therefore, such compounds are useful for treating pulmonary disorders, such as chronic obstructive pulmonary disease and asthma.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional application of U.S. application Ser.No. 10/813,745, filed Mar. 31, 2004 (now U.S. Pat. No. 7,317,102 B2);which application claims the benefit of U.S. Provisional Application No.60/459,291, filed on Apr. 1, 2003; the entire disclosure of which isincorporated herein by reference in its entirety.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to novel diarylmethyl and relatedcompounds having both β₂ adrenergic receptor agonist and muscarinicreceptor antagonist activity. This invention also relates topharmaceutical compositions comprising such compounds, processes andintermediates for preparing such compounds and methods of using suchcompounds to treat pulmonary disorders.

2. State of the Art

Pulmonary disorders, such as asthma and chronic obstructive pulmonarydisease (COPD), are commonly treated with bronchodilators. One class ofbronchodilator in widespread use consists of β₂ adrenergic receptor(adrenoceptor) agonists, such as albuterol, formoterol and salmeterol.These compounds are generally administered by inhalation. Another classof bronchodilator consists of muscarinic receptor antagonists(anticholinergic compounds), such as ipratropium and tiotropium. Thesecompounds are also typically administered by inhalation.

Pharmaceutical compositions containing both a β₂ adrenergic receptoragonist and a muscarinic receptor antagonist are also known in the artfor use in treating pulmonary disorders. For example, U.S. Pat. No.6,433,027 discloses medicament compositions containing a muscarinicreceptor antagonist, such as tiotropium bromide, and a β₂ adrenergicreceptor agonist, such as formoterol fumarate.

Although compounds having either β₂ adrenergic receptor agonist ormuscarinic receptor antagonist activity are known, no compound havingboth β₂ adrenergic receptor agonist and muscarinic receptor antagonistactivity has been previously disclosed. Compounds possessing both β₂adrenergic receptor agonist and muscarinic receptor antagonist activityare highly desirable since such bifunctional compounds would providebronchodilation through two independent modes of action while havingsingle molecule pharmacokinetics.

SUMMARY OF THE INVENTION

The present invention provides novel diarylmethyl and related compoundsthat are useful for treating pulmonary disorders. Among otherproperties, compounds of this invention have been found to possess bothβ₂ adrenergic receptor agonist and muscarinic receptor antagonistactivity.

Accordingly, in one of its composition aspects, the present invention isdirected to a compound of formula I:

wherein:

Ar¹ represents phenyl, (3-6C)cycloalkyl, (3-5C)heteroaryl or(3-5C)heterocyclyl; wherein the heteroaryl and heterocyclyl groupscontain 1 or 2 ring heteroatoms selected independently from oxygen,nitrogen and sulfur;

a is 0 or an integer from 1 to 3;

each R¹ is selected independently from (1-4C)alkyl, (2-4C)alkenyl,(2-4C)alkynyl, (3-6C)cycloalkyl, cyano, halo, —OR^(1a), —SR^(1b),—S(O)R^(1c), —S(O)₂R^(1d), —NR^(1e)R^(1f) and —C(O)OR^(1g); or twoadjacent R¹ groups are joined together to form (3-6C)alkylene,(2-4C)alkylene-O— or —O-(1-4C)alkylene)-O—;

each of R^(1a), R^(1b), R^(1c), R^(1d), R^(1e), R^(1f) and R^(1g) isindependently hydrogen, (1-4C)alkyl or phenyl-(1-4C)alkyl;

Ar² represents phenyl, (3-6C)cycloalkyl, (3-5C)heteroaryl or(3-5C)heterocyclyl; wherein the heteroaryl and heterocyclyl groupcontain 1 or 2 ring heteroatoms selected independently from oxygen,nitrogen and sulfur;

b is 0 or an integer of from 1 to 3;

each R² is selected independently from (1-4C)alkyl, (2-4C)alkenyl,(2-4C)alkynyl, (3-6C)cycloalkyl, cyano, halo, —OR^(2a), —SR^(2b),S(O)R^(2c), —S(O)₂R^(2d), NR^(2e)R^(2f) and —C(O)OR^(2g); or twoadjacent R² groups are joined together to form (3-6C)alkylene,(2-4C)alkylene-O— or —O-(1-4C)alkylene)-O—;

each of R^(2a), R^(2b), R^(2c), R^(2d), R^(2e), R^(2f) and R^(2g) isindependently hydrogen, (1-4C)alkyl or phenyl-(1-4C)alkyl;

E is —CN, —OH, —C(O)NW^(a)W^(b) or —C(O)OW^(c);

W^(a) and W^(b) are selected independently from hydrogen, (1-4C)alkyl orphenyl-(1-4C)alkyl, or together with the nitrogen atom to which they areattached, W^(a) and W^(b) form a pyrrolidin-1-yl, piperidin-1-yl,piperazin-1-yl, morpholin-4-yl or thiomorpholin-4-yl group; or W^(a) andone R¹ are joined to form a covalent bond;

W^(c) is hydrogen, (1-4C)alkyl or phenyl-(1-4C)alkyl;

c is 0 or an integer of from 1 to 4;

each R³ is independently selected from (1-4C)alkyl, (2-4C)alkenyl,(2-4C)alkynyl, (3-6C)cycloalkyl, cyano, halo, —OR^(3a), —SR^(3b),—S(O)R^(3c), S(O)₂R^(3d) and —NR^(3e)R^(3f) and —C(O)OR^(3g); or two R³groups are joined to form (1-3C)alkylene, (2-3C)alkenylene oroxiran-2,3-diyl;

each of R^(3a), R^(3b), R^(3c), R^(3d), R^(3e), R^(3f) and R^(3g) isindependently hydrogen, (1-4C)alkyl or phenyl-(1-4C)alkyl;

z is 1 or 2;

R⁴ is a divalent group of the formula:—(R^(4a))_(d)-(A¹)_(e)-(R^(4b))_(f)-Q-(R^(4c))_(g)-(A²)_(h)-(R^(4d))_(i)—wherein

d, e, f, g, h and i are each independently selected from 0 and 1;

R^(4a), R^(4b), R^(4c) and R^(4d) are each independently selected from(1-10C)alkylene, (2-10C)alkenylene and (2-10C)alkynylene, wherein eachalkylene, alkenylene or alkynylene group is unsubstituted or substitutedwith from 1 to 5 substituents independently selected from (1-4C)alkyl,fluoro, hydroxy, phenyl and phenyl-(1-4C)alkyl;

A¹ and A² are each independently selected from (3-7C)cycloalkylene,(6-10C)arylene, —O-(6-10C)arylene, (6-10C)arylene-O—,(2-9C)heteroarylene, —O-(2-9C)heteroarylene, (2-9C)heteroarylene-O— and(3-6C)heterocyclene, wherein each cycloalkylene is unsubstituted orsubstituted with from 1 to 4 substitutents selected independently from(1-4C)alkyl, and each arylene, heteroarylene or heterocyclene group isunsubstituted or substituted with from 1 to 4 substituents independentlyselected from halo, (1-4C)alkyl, (1-4C)alkoxy, —S-(1-4C)alkyl,—S(O)-(1-4C)alkyl, —S(O)₂-(1-4C)alkyl, —C(O)O(1-4C)alkyl, carboxy,cyano, hydroxy, nitro, trifluoromethyl and trifluoromethoxy;

Q is selected from a bond, —O—, —C(O)O—, —OC(O)—, —S—, —S(O)—, —S(O)₂—,—N(Q^(a))C(O)—, —C(O)N(Q^(b))-, —N(Q^(c))S(O)₂—, —S(O)₂N(Q^(d))-,—N(Q^(e))C(O)N(Q^(f))-, —N(Q^(g))S(O)₂N(Q^(h))-, —OC(O)N(Q^(i))-,—N(Q^(j))C(O)O— and —N(Q^(k));

Q^(a), Q^(b), Q^(c), Q^(d), Q^(e), Q^(f), Q^(g), Q^(h), Q^(i), Q^(j) andQ^(k) are each independently selected from hydrogen, (1-6C)alkyl, A³ and(1-4C)alkylene-A⁴, wherein the alkyl group is unsubstituted orsubstituted with from 1 to 3 substituents independently selected fromfluoro, hydroxy and (1-4C)alkoxy; or together with the nitrogen atom andthe group R^(4b) or R^(4c) to which they are attached, form a 4-6membered azacycloalkylene group;

A³ and A⁴ are each independently selected from (3-6C)cycloalkyl,(6-10C)aryl, (2-9C)heteroaryl and (3-6C)heterocyclyl, wherein eachcycloalkyl is unsubstituted or substituted with from 1 to 4substitutents selected independently from (1-4C)alkyl and each aryl,heteroaryl or heterocyclyl group is unsubstituted or substituted withfrom 1 to 4 substituents independently selected from halo, (1-4C)alkyland (1-4C)alkoxy;

provided that the number of contiguous atoms in the shortest chainbetween the two nitrogen atoms to which R⁴ is attached is in the rangeof from 4 to 16;

R⁵ represents hydrogen or (1-4C)alkyl;

R⁶ is —NR^(6a)CR^(6b)(O) or —CR^(6c)R^(6d)OR^(6e) and R⁷ is hydrogen; orR⁶ and R⁷ together form —NR^(7a)C(O)—CR^(7b)═CR^(7c)—,—CR^(7d)═CR^(7e)—C(O)NR^(7f)—, —NR^(7g)C(O)—CR^(7h)R^(7i)—CR^(7j)R^(7k)—or —CR^(7l)R^(7m)—CR^(7n)R^(7o)— C(O)—NR^(7p)—;

each of R^(6a), R^(6b), R^(6c), R^(6d) and R^(6e) is independentlyhydrogen or (1-4C)alkyl; and

each of R^(7a), R^(7b), R^(7c), R^(7d), R^(7e), R^(7f), R^(7g), R^(7h),R^(7i), R^(7j), R^(7k), R^(7l), R^(7m), R^(7n), R^(7o) and R^(7p) isindependently hydrogen or (1-4C)alkyl;

wherein each alkyl, alkenyl, alkylene and cycloalkyl group in R¹,R^(1a-g), R², R^(2a-g), R³, R^(3a-g), W^(a-c) is optionally substitutedwith from 1 to 5 fluoro substituents;

or a pharmaceutically acceptable salt or solvate or stereoisomerthereof.

In another of its composition aspects, this invention is directed to acompound of formula II:

wherein

R⁴, R⁶ and R⁷ are as defined herein (including any specific or preferredembodiments);

or a pharmaceutically acceptable salt or solvate or stereoisomerthereof.

In another of its composition aspects, this invention is directed to acompound of formula III:

wherein

R⁴ is as defined herein (including any specific or preferredembodiments);

or a pharmaceutically acceptable salt or solvate or stereoisomerthereof.

In yet another of its composition aspects, this invention is directed toa compound of formula IV:

wherein

R⁴ is as defined herein (including any specific or preferredembodiments);

or a pharmaceutically acceptable salt or solvate or stereoisomerthereof.

In still another of its composition aspects, this invention is directedto a compound of formula V:

wherein

R⁴ is as defined herein (including any specific or preferredembodiments);

or a pharmaceutically acceptable salt or solvate or stereoisomerthereof.

In another of its composition aspects, this invention is directed to apharmaceutical composition comprising a pharmaceutically acceptablecarrier and a therapeutically effective amount of a compound of formulaI or a pharmaceutically acceptable salt or solvate or stereoisomerthereof. Such pharmaceutical compositions may optionally contain othertherapeutic agents. Accordingly, in one embodiment, this invention isdirected to such a pharmaceutical composition wherein the compositionfurther comprises a therapeutically effective amount of a steroidalanti-inflammatory agent, such as a corticosteroid.

Compounds of this invention possess both β₂ adrenergic receptor agonistactivity and muscarinic receptor antagonist activity. Accordingly, thecompounds of formula I are useful for treating pulmonary disorders, suchas asthma and chronic obstructive pulmonary disease.

Accordingly, in one of its method aspects, this invention is directed toa method for treating a pulmonary disorder, the method comprisingadministering to a patient in need of treatment a therapeuticallyeffective amount of a compound of formula I or a pharmaceuticallyacceptable salt or solvate or stereoisomer thereof.

Additionally, in another of its method aspects, this invention isdirected to a method of providing bronchodilation in a patient, themethod comprising administering to a patient requiring bronchodilation atherapeutically effective amount of a compound of formula I or apharmaceutically acceptable salt or solvate or stereoisomer thereof.

This invention is also directed to a method of treating chronicobstructive pulmonary disease or asthma, the method comprisingadministering to a patient in need of treatment a therapeuticallyeffective amount of a compound of formula I or a pharmaceuticallyacceptable salt or solvate or stereoisomer thereof.

Since compounds of this invention possess both β₂ adrenergic receptoragonist activity and muscarinic receptor antagonist activity, suchcompounds are also useful as research tools. Accordingly, in yet anotherof its method aspects, this invention is directed to a method for usinga compound of formula I or a pharmaceutically acceptable salt or solvateor stereoisomer thereof as a research tool for studying a biologicalsystem or sample, or for discovering new chemical compounds having bothβ₂ adrenergic agonist activity and muscarinic receptor antagonistactivity.

This invention is also directed to processes and novel intermediatesuseful for preparing compounds of formula I or a pharmaceuticallyacceptable salt or solvate or stereoisomer thereof. Accordingly, inanother of its method aspects, this invention is directed to a processof preparing a compound of formula 1, the process comprising:

(a) reacting a compound of formula 1 or a salt thereof, with a compoundof formula 2;

(b) reacting a compound of formula 3 or a salt thereof, with a compoundof formula 4;

(c) coupling a compound of formula 5 with a compound of formula 6;

(d) for a compound of formula I wherein R⁵ represents a hydrogen atom,reacting a compound of formula 3 with a compound of formula 7 or ahydrate thereof, in the presence of a reducing agent;

(e) reacting a compound of formula 1 with a compound of formula 8 or ahydrate thereof, in the presence of a reducing agent;

(f) reacting a compound of formula 9, with a compound of formula 10;

(g) reacting a compound of formula 11 with a reducing agent;

(h) for a compound of formula I in which E represents —C(O)NW^(a)W^(b),reacting a compound of formula 12 with a compound of formula 13; or

(i) reacting a compound of formula 14 or a hydrate thereof, with acompound of formula 10 in the presence of a reducing agent;

and then removing any protecting groups to form a compound of formula I;wherein the compounds of formula 1-14 are as defined therein.

In one embodiment, the above process further comprises the step offorming a pharmaceutically acceptable salt of a compound of formula 1.In other embodiments, this invention is directed to the other processesdescribed herein; and to the product prepared by any of the processesdescribed herein.

This invention is also directed to a compound of formula I or apharmaceutically acceptable salt or solvate or stereoisomer thereof, foruse in therapy or as a medicament.

Additionally, this invention is directed to the use of a compound offormula I or a pharmaceutically acceptable salt or solvate orstereoisomer thereof, for the manufacture of a medicament; especiallyfor the manufacture of a medicament for the treatment of a pulmonarydisorder.

DETAILED DESCRIPTION OF THE INVENTION

In one of its composition aspects, this invention is directed to novelcompounds of formula I or pharmaceutically acceptable salts or solvatesor stereoisomers thereof. These compounds contain one or more chiralcenters and therefore, this invention is directed to racemic mixtures;pure stereoisomers (i.e., enantiomers or diastereomers);stereoisomer-enriched mixtures and the like unless otherwise indicated.When a particular stereoisomer is shown or named herein, it will beunderstood by those skilled in the art that minor amounts of otherstereoisomers may be present in the compositions of this inventionunless otherwise indicated, provided that the utility of the compositionas a whole is not eliminated by the presence of such other isomers.

In particular, compounds of formula I contain a chiral center at thecarbon atom indicated by the symbol * in the following formula:

In one embodiment of this invention, the carbon atom identified by thesymbol * has the (R) configuration. In this embodiment, it is preferredfor compounds of formula I to have the (R) configuration at the carbonatom identified by the symbol * or to be enriched in a stereoisomericform having the (R) configuration at this carbon atom. In anotherembodiment of this invention, the carbon atom identified by the symbol *has the (S) configuration. In this embodiment, it is preferred forcompounds of formula I to have the (S) configuration at the carbon atomidentified by the symbol * or to be enriched in a stereoisomeric formhaving the (S) configuration at this carbon atom. In some cases, inorder to optimize the β₂ adrenergic agonist activity of the compounds ofthis invention, it is preferred that the carbon atom identified by thesymbol * has the (R) configuration.

Additionally, when z is 1, the compound of formula I contain a chiralcenter at the carbon atom indicated by the symbol ** in the followingformula:

In one embodiment of this invention, the carbon atom identified by thesymbol ** has the (S) configuration. In this embodiment, it is preferredfor compounds of formula I to have the (S) configuration at the carbonatom identified by the symbol ** or to be enriched in a stereoisomericform having the (S) configuration at this carbon atom. In anotherembodiment of this invention, the carbon atom identified by the symbol** has the (R) configuration. In this embodiment, it is preferred forcompounds of formula I to have the (R) configuration at the carbon atomidentified by the symbol ** or to be enriched in a stereoisomeric formhaving the (R) configuration at this carbon atom.

The compounds of formula I also contain several basic groups (e.g.,amino groups) and therefore, the compounds of formula I can exist as thefree base or in various salt forms. All such salt forms are includedwithin the scope of this invention. Furthermore, solvates of compoundsof formula I or salts thereof are included within the scope of thisinvention.

Additionally, where applicable, all cis-trans or E/Z isomers (geometricisomers), tautomeric forms and topoisomeric forms of the compounds offormula I are included within the scope of this invention unlessotherwise specified.

The nomenclature used herein to name the compounds of this invention andintermediates thereof has generally been derived using thecommercially-available AutoNom software (MDL, San Leandro, Calif.).

Representative Embodiments

The following substituents and values are intended to providerepresentative examples of various aspects and embodiments of thisinvention. These representative values are intended to further defineand illustrate such aspects and embodiments and are not intended toexclude other embodiments or to limit the scope of this invention. Inthis regard, the representation that a particular value or substituentis preferred is not intended in any way to exclude other values orsubstituents from this invention unless specifically indicated.

In one embodiment, Ar¹ represents phenyl optionally substituted with 1to 3 R¹ groups. When Ar¹ is phenyl, R¹ may be at the 2, 3, 4, 5 or6-position of the phenyl ring. Representative Ar¹ groups in thisembodiment include phenyl, 2-chlorophenyl, 3-chlorophenyl,4-chlorophenyl, 2-fluorophenyl, 3-fluorophenyl, 4-fluorophenyl,2-methylphenyl, 3-methylphenyl, 4-methylphenyl, 3,4-difluorophenyl,3-chloro-4-fluorophenyl, and the like.

In another embodiment, Ar¹ represents (3-6C)cycloalkyl optionallysubstituted with 1 to 3 R¹ groups. When Ar¹ is cycloalkyl, R¹ may be atany available position of the cycloalkyl group. Representative Ar¹groups in this embodiment include cyclopropyl, cyclobutyl, cyclopentyland cyclohexyl.

In another embodiment, Ar¹ represents (3-5C)heteroaryl optionallysubstituted with 1 to 3 R¹ groups. When Ar¹ is heteroaryl, R¹ may be atany available position of the heteroaryl group. Representative Ar¹groups in this embodiment include pyridyl, such as pyrid-2-yl,pyrid-3-yl or pyrid-4-yl; furyl, such as fur-2-yl and fur-3-yl; thienyl,such as thien-2-yl and thien-3-yl; and a pyrrolyl, such as pyrrol-2-yland pyrrol-3-yl.

In still another embodiment, Ar¹ represents (3-5C)heterocyclyloptionally substituted with 1 to 3 R¹ groups. When Ar¹ is heterocyclyl,R¹ may be at any available position of the heterocyclyl group.Representative Ar¹ groups in this embodiment include piperidinyl, suchas piperidin-4-yl; and pyrrolidinyl, such as pyrrolidin-2-yl.

In a particular embodiment, Ar¹ is optionally substituted phenyl,pyridyl, thienyl, cyclobutyl, cyclopentyl or cyclohexyl.

In one embodiment, Ar² represents phenyl optionally substituted with 1to 3 R² groups. When Ar² is phenyl, R² may be at the 2, 3, 4, 5 or6-position of the phenyl ring. Representative Ar² groups in thisembodiment include phenyl, 2-chlorophenyl, 3-chlorophenyl,4-chlorophenyl, 2-fluorophenyl, 3-fluorophenyl, 4-fluorophenyl,2-methylphenyl, 3-methylphenyl, 4-methylphenyl, 3,4-difluorophenyl,3-chloro-4-fluorophenyl, and the like.

In another embodiment, Ar² represents (3-6C)cycloalkyl optionallysubstituted with 1 to 3 R² groups. When Ar² is cycloalkyl, R² may be atany available position of the cycloalkyl group. Representative Ar²groups in this embodiment include cyclopropyl, cyclobutyl, cyclopentyland cyclohexyl.

In another embodiment, Ar² represents (3-5C)heteroaryl optionallysubstituted with 1 to 3 R² groups. When Ar² is heteroaryl, R² may be atany available position of the heteroaryl group. Representative Ar²groups in this embodiment include pyridyl, such as pyrid-2-yl,pyrid-3-yl or pyrid-4-yl; furyl, such as fur-2-yl and fur-3-yl; thienyl,such as thien-2-yl and thien-3-yl; and a pyrrolyl, such as pyrrol-2-yland pyrrol-3-yl.

In still another embodiment, Ar² represents (3-5C)heterocyclyloptionally substituted with 1 to 3 R² groups. When Ar² is heterocyclyl,R² may be at any available position of the heterocyclyl group.Representative Ar² groups in this embodiment include piperidinyl, suchas piperidin-4-yl; and pyrrolidinyl, such as pyrrolidin-2-yl.

In a particular embodiment, Ar² is optionally substituted phenyl,pyridyl, thienyl, cyclobutyl, cyclopentyl, cyclohexyl or piperidinyl.

In a particular embodiment, Ar¹ is optionally substituted phenyl and Ar²is optionally substituted phenyl, pyridyl, thienyl, cyclobutyl,cyclopentyl or cyclohexyl.

In particular embodiments of the compounds of formula I, a and b areindependently 0, 1 or 2; including 0 or 1. In one embodiment, both a andb are 0.

When present, each R¹ may be at any available position of the Ar¹ ringto which it is attached. In one embodiment, each R¹ is independentlyselected from (1-4C)alkyl, halo, —OR^(1a) and —NR^(1f)R^(1g); whereinthe alkyl group is optionally substituted with 1 to 3 fluorosubstituents; such as methyl, fluoro, chloro, bromo, hydroxy, methoxy,amino, methylamino, dimethylamino, trifluoromethyl and the like.Particular values for R¹ are fluoro or chloro.

When present, each R² may be at any available position of the Ar² ringto which it is attached. In one embodiment, each R² is independentlyselected from (1-4C)alkyl, halo, —OR^(2a) and —NR^(2f)R^(2g), whereinthe alkyl group is optionally substituted with 1 to 3 fluorosubstituents; such as methyl, fluoro, chloro, bromo, hydroxy, methoxy,amino, methylamino, dimethylamino, trifluoromethyl and the like.Particular values for R² are fluoro or chloro.

Each R^(1a), R^(1b), R^(1c), R^(1d), R^(1e), R^(1f) and R^(1g) andR^(2a), R^(2b), R^(2c), R^(2d), R^(2e), R^(2f) and R^(2g) as used in R¹and R², respectively, is independently hydrogen, (1-4C)alkyl orphenyl-(1-4C)alkyl; wherein the alkyl group is optionally substitutedwith 1 to 3 fluoro substituents; such as hydrogen, methyl,trifluoromethyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl,isobutyl, tert-butyl or benzyl. In one embodiment, these groups areindependently hydrogen or (1-3C)alkyl. In another embodiment, thesegroups are independently hydrogen, methyl or ethyl.

In one embodiment, E is —C(O)NW^(a)W^(b). In another embodiment, E is—CN.

In a particular embodiment, W^(a) and W^(b) are independently hydrogenor (1-4C)alkyl; such as hydrogen, methyl, ethyl, n-propyl, isopropyl,n-butyl, sec-butyl, isobutyl and tert-butyl. In one embodiment, W^(a)and W^(b) are independently hydrogen or (1-3C)alkyl. In anotherembodiment, W^(a) and W^(b) are independently hydrogen, methyl or ethyl;such as hydrogen or methyl. In yet another embodiment, W^(a) and W^(b)are both hydrogen and E is —C(O)NH₂.

In another particular embodiment, when Ar¹ is phenyl, W^(a) is joinedwith one R¹ group at the 2-position of the phenyl ring to form acovalent bond. In this embodiment, W^(a) and R¹ form an indolone moietyof the formula:

wherein the remainder of the compound is as shown in formula I.

In a particular embodiment of the compounds of formula I, c is 0, 1 or2; including 0 or 1. In one embodiment, c is 0.

In one embodiment, each R³ is independently selected from (1-4C)alkyl;wherein the alkyl group is optionally substituted with 1 to 3 fluorosubstituents; such as methyl, trifluoromethyl, ethyl, n-propyl,isopropyl, n-butyl, sec-butyl, isobutyl and tert-butyl. In anotheraspect, each R³ is independently methyl or ethyl.

In yet another embodiment, two R³ groups are joined to form a(1-3C)alkylene or (2-3C)alkenylene group. For example, when z is 2, twoR³ groups at the 2 and 6-positions on the piperidine ring can be joinedto form an ethylene bridge (i.e., the piperidine ring and the R³ groupsform an 8-azabicyclo[3.2.1]octane ring); or two R³ groups at the 1 and4-positions on the piperidine ring can be joined to form an ethylenebridge (i.e., the piperidine ring and the R³ groups form an1-azabicyclo[2.2.2]octane ring). In this embodiment, other R³ groups asdefined herein may also be present.

In still another embodiment, two R³ groups are joined to form aoxiran-2,3-diyl group. For example, when z is 2, two R³ groups at the 2and 6-positions on the piperidine ring can be joined to form a3-oxatricyclo[3.3.1.0^(2.4)]nonane ring). In this embodiment, other R³groups as defined herein may also be present.

Each R^(3a), R^(3b), R^(3c), R^(3d), R^(3e), R^(3f) and R^(3g) as usedin R³ is independently hydrogen or (1-4C)alkyl; wherein the alkyl groupis optionally substituted with 1 to 3 fluoro substituents; such ashydrogen, methyl, trifluoromethyl, ethyl, n-propyl, isopropyl, n-butyl,sec-butyl, isobutyl and tert-butyl. In one embodiment, these groups areindependently hydrogen or (1-3C)alkyl. In another embodiment, thesegroups are independently hydrogen, methyl or ethyl.

By way of further illustration, in separate and distinct particularembodiments of the present invention, the group:

is selected from:

wherein R¹, R², R³, W^(a), W^(b), a, b and c are as defined herein.

In one embodiment of the compounds of formula I, R⁵ is hydrogen or(1-4C)alkyl; such as hydrogen, methyl, ethyl, n-propyl, isopropyl,n-butyl, sec-butyl, isobutyl and tert-butyl. In another embodiment, eachR⁵ is independently hydrogen, methyl or ethyl. In a particularembodiment, R⁵ is hydrogen.

In one embodiment of this invention, R⁶ is —NR^(6a)CR^(6b)(O) and R⁷ ishydrogen, where each of R^(6a) and R^(6b) is independently hydrogen or(1-4C)alkyl, such as hydrogen, methyl, ethyl, n-propyl, isopropyl,n-butyl, sec-butyl, isobutyl and tert-butyl. In one embodiment, thesegroups are independently hydrogen or (1-3C)alkyl. In another embodiment,these groups are independently hydrogen, methyl or ethyl. A particularvalue for R⁶ in this embodiment is —NHCHO.

In another embodiment, R⁶ and R⁷ together form—NR^(7a)C(O)—CR^(7b)═CR^(7c)—, —CR^(7d)═CR^(7e)—C(O)—NR^(7f)—,—NR^(7g)C(O)—CR^(7h)R^(7i)—CR^(7j)R^(7k) or—CR^(7l)R^(7m)—CR^(7n)R^(7o)—C(O) —NR^(7p)—; where each of R^(7a),R^(7b), R^(7c), R^(7d), R^(7e), R^(7f), R^(7g), R^(7h), R^(7i), R^(7j),R^(7k), R^(7l), R^(7m), R^(7n), R^(7o) and R^(7p) is independentlyhydrogen or (1-4C)alkyl; such as hydrogen, methyl, ethyl, n-propyl,isopropyl, n-butyl, sec-butyl, isobutyl and tert-butyl. In oneembodiment, these groups are independently hydrogen or (1-3C)alkyl. Inanother embodiment, these groups are independently hydrogen, methyl orethyl. Particular values for R⁶ and R⁷ in this embodiment are R⁶ and R⁷together form —NHC(O)—CH═CH—, —CH═CH—C(O)—NH—, —CH₂—CH₂—C(O)NH— or—NHC(O)—CH₂—CH₂—; including where R⁶ and R⁷ together form —NHC(O)—CH═CH—or —CH═CH—C(O)—NH—; and in particular, where R⁶ and R⁷ together form—NHC(O)—CH═CH— (i.e., the nitrogen atom is attached at R⁶ and the carbonatom is attached at R⁷ to form, together with the hydroxyphenyl ring towhich R⁶ and R⁷ are attached, a 8-hydroxy-2-oxo-1,2-dihydroquinolin-5-ylgroup).

In the compounds of formula I, R⁴ is a divalent group of the formula:—(R^(4a))_(d)-(A¹)_(e)-(R^(4b))_(f)-Q-(R^(4c))_(g)-(A²)_(h)-(R^(4d))_(i)—

wherein R^(4a), A¹, R^(4b), Q, R^(4c), A, R^(4d), d, e, f, g h and i areas defined herein. In the compound of this invention, the values of eachof the components R^(4a), A¹, R^(4b)b, Q, R^(4c), A² and R^(4d) areselected such that the number of contiguous atoms in the shortest chainbetween the two nitrogen atoms to which R⁴ is attached is in the rangeof from 4 to 16, (specifically, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15or 16); including 8, 9, 10, 11, 12, 13 or 14; such as 8, 9, 10 or 11; or9 or 10. When selecting values for each variable in R⁴, it will beappreciated by those skilled in the art that values should be selectedsuch that a chemically stable group is formed.

When determining the number of contiguous atoms in the shortest chainbetween the two nitrogen atoms to which R⁴ is attached, each contiguousatom of the chain is counted consecutively starting from the first atomin the R⁴ group adjacent to the nitrogen of the piperidine ring endingwith the last atom in the R⁴ group adjacent to the nitrogen of theaminohydroxyethyl group. Where two or more chains are possible, theshortest chain is used to determine the number of contiguous atoms. Asshown below, for example, when R⁴ is—(CH₂)₂—NHC(O)—CH₂-(phen-1,4-ylene)-CH₂—, there are 10 contiguous atomsin the shortest chain counted consecutively starting from the first atomin the R⁴ group adjacent to the nitrogen of the piperidine ring endingwith the last atom in the R⁴ group adjacent to the nitrogen of theaminohydroxyethyl group as shown below:

In one embodiment of R⁴, R^(4a) is selected from (1-10C)alkylene,(2-10C)alkenylene and (2-10C)alkynylene wherein the alkylene group isunsubstituted or substituted with 1 or 2 substituents independentlyselected from (1-4C)alkyl, hydroxy and phenyl. Representative examplesof particular values for R^(4a) are —(CH₂)₂—, —(CH₂)₃—, —(CH₂)₄—,—(CH₂)₅—, —(CH₂)₆—, —(CH₂)₇—, —(CH₂)₈—, —(CH₂)₉—, —(CH₂)₁₀—,—(CH₂)CH(CH₃)—, —(CH₂)C(CH₃)₂—, and —(CH₂)₂C(phenyl)₂-. In anotheraspect, R^(4a) is —(CH₂)C(═CH₂)—.

In one embodiment, d is 1.

In one embodiment, A¹ is an optionally substituted (3-7C)cycloalkylenegroup; including a cyclohexylene group, such as cyclohex-1,4-ylene andcyclohex-1,3-ylene; and a cyclopentylene group, such ascyclopent-1,3-ylene.

In another embodiment, A¹ is an optionally substituted (6-10C)arylenegroup, including a phenylene group, such as phen-1,4-ylene,phen-1,3-ylene and phen-1,2-ylene; and a naphthylene group, such asnaphth-1,4-ylene and napth-1,5-ylene.

In yet another embodiment, A¹ is an optionally substituted(2-9C)heteroarylene group, including a pyridylene group, such aspyrid-1,4-ylene; a furylene group, such as fur-2,5-ylene andfur-2,4-ylene; a thienylene group, such as thien-2,5-ylene andthien-2,4-ylene; and a pyrrolylene, such as pyrrol-2,5-ylene andpyrrol-2,4-ylene.

In still another embodiment, A¹ is an optionally substituted(3-6C)heterocyclene group, including a piperidinylene group, such aspiperidin-1,4-ylene; and a pyrrolidinylene group, such aspyrrolidin-2,5-ylene.

In a particular embodiment, A¹ is an optionally substituted phenylene,thienylene, cyclopentylene, cyclohexylene or piperidinylene.

In one embodiment, e is 0.

In a particular embodiment, R^(4b) is (1-5C)alkylene. Representativeexamples of particular values for R^(4b) are —CH₂—, —(CH₂)₂—, —(CH₂)₃—,—(CH₂)₄—, —(CH₂)₅—; including methylene, ethylene and propylene.

In one embodiment, f is 0.

In a particular embodiment, Q is selected from a bond, —N(Q^(a))C(O)—,—C(O)N(Q^(b))-, —N(Q^(c))S(O)₂—, —S(O)₂N(Q^(d))-,—N(Q^(e))C(O)N(Q^(f))-, —OC(O)N(Q^(i))-, —N(Q^(i))C(O)O— or —N(Q^(k));such as where Q is a bond, —N(Q^(a))C(O)— or —C(O)N(Q^(b))-.Representative examples of particular values for Q are a bond, O, NH,—C(O)NH—, —C(O)N(CH₃)—, —NHC(O)—, —N(CH₃)C(O)—, —S(O)₂NH—,—S(O)₂N(CH₃)—, —NHS(O)₂—, —N(CH₃)S(O)₂— and —NHC(O)NH—. Another exampleof a value for Q, together with R^(4c), is —C(O)(piperidin-1,4-ylene).

In one embodiment, Q^(a), Q^(b), Q^(c), Q^(d), Q^(e), Q^(f), Q^(g),Q^(h), Q^(i), Q^(j) and Q^(k) are each independently selected fromhydrogen and (1-6C)alkyl, wherein the alkyl group is unsubstituted orsubstituted with from 1 to 3 substituents independently selected fromfluoro, hydroxy and (1-4C)alkoxy. For example, Q^(a), Q^(b), Q^(c),Q^(d), Q^(e), Q^(f), Q^(g), Q^(h), Q^(i), Q^(j) and Q^(k) are eachindependently selected from hydrogen, and (1-3C)alkyl, includinghydrogen, methyl, ethyl, n-propyl and isopropyl. An example of a valuefor each of Q^(a), Q^(b), Q^(c), Q^(d), Q^(e), Q^(f), Q^(g), Q^(h),Q^(i), Q^(j) and Q^(k) is hydrogen.

In another embodiment, Q^(a), Q^(b), Q^(c), Q^(d), Q^(e), Q^(f), Q^(g),Q^(h), Q^(i), Q^(j) and Q^(k) together with the nitrogen atom and thegroup R^(4b) or R^(4c) to which they are attached, form a 4-6 memberedazacycloalkylene group. For example, Q^(a) and Q^(b) together with thenitrogen atom and the group R^(4b) or R^(4c) to which they are attached,form a piperidin-4-ylene group. By way of illustration, when Qrepresents —N(Q^(a))C(O)— and Q^(a) together with the nitrogen atom andthe group R^(4b) to which it is attached, forms a piperidin-4-ylenegroup, R⁴ is a group of formula:

Similarly, when Q represents —C(O)N(Q^(b))- and Q^(b) together with thenitrogen atom and the group R^(4c) to which it is attached, forms apiperidin-4-ylene group, R⁴ is a group of formula:

In a particular embodiment, R^(4c) is (1-5C)alkylene. Representativeexamples of particular values for R^(4c) are —CH₂—, —(CH₂)₂—, —(CH₂)₃—,—(CH₂)₄—, —(CH₂)₅—; including methylene, ethylene and propylene.

In one embodiment, A² is an optionally substituted (3-7C)cycloalkylenegroup; including a cyclohexylene group, such as cyclohex-1,4-ylene andcyclohex-1,3-ylene; and a cyclopentylene group, such ascyclopent-1,3-ylene.

In another embodiment, A² is an optionally substituted (6-10C)arylenegroup, including a phenylene group, such as phen-1,4-ylene,phen-1,3-ylene and phen-1,2-ylene; and a naphthylene group, such asnaphth-1,4-ylene and napth-1,5-ylene.

In yet another embodiment, A² is an optionally substituted(2-9C)heteroarylene group, including a pyridylene group, such aspyrid-1,4-ylene; a furylene group, such as fur-2,5-ylene andfur-2,4-ylene; a thienylene group, such as thien-2,5-ylene andthien-2,4-ylene; and a pyrrolylene, such as pyrrol-2,5-ylene andpyrrol-2,4-ylene.

In still another embodiment, A² is an optionally substituted(3-6C)heterocyclene group, including a piperidinylene group, such aspiperidin-1,4-ylene; and a pyrrolidinylene group, such aspyrrolidin-2,5-ylene.

In a particular embodiment, A² is optionally substituted phenylene,thienylene, cyclopentylene, cyclohexylene or piperidinylene.

By way of illustration, either A¹ or A² or both can be phenylene, suchas phen-1,4-ylene or phen-1,3-ylene, where the phenylene group isunsubstituted or substituted with from 1 to 4 substituents independentlyselected from halo, (1-4C)alkyl, (1-4C)alkoxy, —S—(1-4C)alkyl,—S(O)—(1-4C)alkyl, —S(O)₂—(1-4C)alkyl, —C(O)O(1-4C)alkyl, carboxy,cyano, hydroxy, nitro, trifluoromethyl and trifluoromethoxy.Representative examples include phen-1,3-ylene, phen-1,4-ylene,4-chlorophen-1,3-ylene, 6-chlorophen-1,3-ylene, 4-methylphen-1,3-ylene,2-fluorophen-1,4-ylene, 2-chlorophen-1,4-ylene, 2-bromophen-1,4-ylene,2-iodophen-1,4-ylene, 2-methylphen-1,4-ylene, 2-methoxyphen-1,4-ylene,2-trifluoromethoxyphen-1,4-ylene, 3-nitrophen-1,4-ylene,3-chlorophen-1,4-ylene, 2,5-difluorophen-1,4-ylene,2,6-dichlorophen-1,4-ylene, 2,6-diiodophen-1,4-ylene,2-chloro-6-methylphen-1,4-ylene, 2-chloro-5-methoxyphen-1,4-ylene,2,3,5,6-tetrafluorophen-1,4-ylene.

Alternatively, A¹ or A² or both can be cyclopentylene or cyclohexylene;wherein the cyclopentylene or cyclohexylene group is unsubstituted orsubstituted with (1-4C)alkyl. Representative examples includecis-cyclopent-1,3-ylene, trans-cyclopent-1,3-ylene,cis-cyclohex-1,4-ylene and trans-cyclohex-1,4-ylene. A¹ or A² or bothcan also be optionally substituted thienylene or piperidinylene, forexample, thien-2,5-ylene or piperidin-1,4-ylene.

In one embodiment, R^(4d) is selected from (1-10C)alkylene,(2-10C)alkenylene and (2-10C)alkynylene wherein the alkylene isunsubstituted or substituted with 1 or 2 substituents independentlyselected from (1-4C)alkyl, hydroxy and phenyl. Representative examplesof particular values for R^(4d) are —(CH₂)—, —(CH₂)₂—, —(CH₂)₃—,—(CH₂)₄—, —(CH₂)₅—, —(CH₂)₆—, —(CH₂)₇—, —(CH₂)₈—, —(CH₂)₉—, —(CH₂)₁₀—and —(CH₂)CH(CH₃)—(CH₂)—C(CH₃)₂—(CH₂)₂—.

In a particular embodiment, R⁴ is a divalent group of the formula:—(R^(4a))_(d)— where R^(4a) is (4-10C)alkylene. In one aspect of thisembodiment, R⁴ is a divalent group of the formula: —(CH₂)_(j)— where jis 8, 9 or 10. Examples of particular values for R⁴ in this embodimentare —(CH₂)₄—, —(CH₂)₅—, —(CH₂)₆—, —(CH₂)₇—, —(CH₂)₈—, —(CH₂)₉, and—(CH₂)₁₀—; including —(CH₂)₈—, —(CH₂)₉, and —(CH₂)₁₀—.

In another particular embodiment, R⁴ is a divalent group of the formula:—(R^(4a))_(d)-(A²)_(h)-(R^(4d))_(i)—

where R^(4a) is (1-10C)alkylene, such as —(CH₂)—, —(CH₂)₂—, —(CH₂)₃—; A²is (6-10C)arylene, such as phen-1,4-ylene or phen-1,3-ylene, or(2-9C)heteroarylene, such as thien-2,5-ylene or thien-2,4-ylene; andR^(4d) is (1-10C)alkylene, such as —(CH₂)—, —(CH₂)₂—, —(CH₂)₃—. Examplesof particular values for R⁴ in this embodiment are—(CH₂)-(phen-1,4-ylene)-(CH₂)—; —(CH₂)-(phen-1,4-ylene)-(CH₂)₂—;—(CH₂)-(phen-1,4-ylene)-(CH₂)₃—; —(CH₂)₂-(phen-1,4-ylene)-(CH₂)—;—(CH₂)₂-(phen-1,4-ylene)-(CH₂)₂—; —(CH₂)₂-(phen-1,4-ylene)-(CH₂)₃—;—(CH₂)₃-(phen-1,4-ylene)-(CH₂)—; —(CH₂)₃-(phen-1,4-ylene)-(CH₂)₂—,—(CH₂)₃-(phen-1,4-ylene)-(CH₂)₃—, —(CH₂)₄-(phen-1,4-ylene)-(CH₂)—;—(CH₂)₄-(phen-1,4-ylene)-(CH₂)₂— and —(CH₂)₄-(phen-1,4-ylene)-(CH₂)₃—.

In yet another particular embodiment, R⁴ is a divalent group of theformula:—(R^(4a))_(d)-Q-(A²)_(h)-(R^(4d))_(i)—

where Q is —O— or —N(Q^(k))-; Q^(k) is hydrogen or (1-3C)alkyl, such asmethyl or ethyl; R^(4a) is (1-10C)alkylene, such as —(CH₂)—, —(CH₂)₂—,—(CH₂)₃—; A² is (6-10C)arylene, such as phen-1,4-ylene orphen-1,3-ylene, or (2-9C)heteroarylene, such as thien-2,5-ylene orthien-2,4-ylene; and R^(4d) is (1-10C)alkylene, such as —(CH₂)—,—(CH₂)₂—, —(CH₂)₃—. Examples of particular values for R⁴ in thisembodiment are —(CH₂)₂—O-(phen-1,4-ylene)-(CH₂)—;—(CH₂)₂—O-(phen-1,4-ylene)-(CH₂)₂—; —(CH₂)₂—O-(phen-1,4-ylene)-(CH₂)₃—;—(CH₂)₃—O-(phen-1,4-ylene)-(CH₂)—; —(CH₂)₃—O-(phen-1,4-ylene)-(CH₂)₂—;—(CH₂)₃—O-(phen-1,4-ylene)-(CH₂)₃—; —(CH₂)₂—NH-(phen-1,4-ylene)-(CH₂)—;—(CH₂)₂—NH-(phen-1,4-ylene)-(CH₂)₂—;—(CH₂)₂—NH-(phen-1,4-ylene)-(CH₂)₃—; —(CH₂)₃—NH-(phen-1,4-ylene)-(CH₂)—;—(CH₂)₃—NH-(phen-1,4-ylene)-(CH₂)₂— and—(CH₂)₃—NH-(phen-1,4-ylene)-(CH₂)₃—.

In yet another particular embodiment, R⁴ is a divalent group of theformula:—(R^(4a))_(d)-(A¹)_(e)-(R^(4b))_(f)-Q-(R^(4c))_(g)-(A²)_(h)-(R^(4d))_(i)—

where Q is —N(Q^(a))C(O)— or —C(O)N(Q^(b))-. A particular value for R⁴in this embodiment is the formula:

where m is an integer from 2 to 10; and n is an integer from 2 to 10;provided that m+n is an integer from 4 to 12. In this formula for R⁴, dand g are 1 and e, f, h and i are 0; and R^(4a) is —(CH₂)_(m)—, R^(4c)is —(CH₂)_(n)— and Q is —C(O)NH—. Particular values for m are 2 or 3;and for n, 4, 5 or 6.

Another particular value for R⁴ is the formula:

where o is an integer from 2 to 7; and p is an integer from 1 to 6;provided that o+p is an integer from 3 to 8. In this formula for R⁴, d,h and i are 1 and e, f and g are 0; and R^(4a) is —(CH₂)_(o)—, A² isphen-1,4-ylene, R^(4d) is —(CH₂)_(p)— and Q is —C(O)NH—. Particularvalues for o are 2 or 3; and for p, 1 or 2. In this embodiment, thephen-1,4-ylene group may be optionally substituted as defined herein forA².

Another particular value for R⁴ is the formula:

where q is an integer from 2 to 6; r is an integer from 1 to 5; and s isan integer from 1 to 5; provided that q+r+s is an integer from 4 to 8.In this formula for R⁴, d, g, h and i are 1 and e and f are 0; andR^(4a) is —(CH₂)_(q)—, R^(4c) is —(CH₂)_(r)—, A² is 1,4-phenylene,R^(4d) is —(CH₂)_(s)— and Q is —C(O)NH—. Particular values for q are 2or 3; for r, 1 or 2; and for s, 1 or 2. In this embodiment, thephen-1,4-ylene group may be optionally substituted as defined herein forA².

Another particular value for R⁴ is the formula:

where t is an integer from 2 to 10; and u is an integer from 2 to 10;provided that t+u is an integer from 4 to 12. In this formula for R⁴, dand g are 1 and e, f, h and i are 0; and R^(4a) is —(CH₂)_(t)—, R^(4c)is —(CH₂)_(u)— and Q is —NHC(O)—. Particular values for t are 2 or 3;and for u, 4, 5 or 6.

Another particular value for R⁴ is the formula:

where v is an integer from 2 to 7; and w is an integer from 1 to 6;provided that v+w is an integer from 3 to 8. In this formula for R⁴, d,h and i are 1 and e, f and g are 0; and R^(4a) is —(CH₂)_(v)—, A² is1,4-phenylene, R^(4d) is —(CH₂)_(w)— and Q is —NHC(O)—. Particularvalues for v are 2 or 3; and for w, 1 or 2. In this embodiment, thephen-1,4-ylene group may be optionally substituted as defined herein forA².

Another particular value for R⁴ is the formula:

where x is an integer from 2 to 6; y is an integer from 1 to 5; and aais an integer from 1 to 5; provided that x+y+aa is an integer from 4 to8. In this formula for R⁴, d, g, h and i are 1 and e and f are 0; andR^(4a) is —(CH₂)_(x)—, R^(4c) is —(CH₂)_(y)—, A² is 1,4-phenylene,R^(4d) is —(CH₂)_(z)— and Q is —NHC(O)—. Particular values for x are 2or 3; for y, 1 or 2; and for aa, 1 or 2. In this embodiment, thephen-1,4-ylene group may be optionally substituted as defined herein forA².

By way of further illustration, R⁴ can be selected from:

-   —(CH₂)₇—;-   —(CH₂)₈—;-   —(CH₂)₉—;-   —(CH₂)₁₀—;-   —(CH₂)₁₁—;-   —(CH₂)₂C(O)NH(CH₂)₅—;-   —(CH₂)₂N(CH₃)C(O)(CH₂)₅—;-   —(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)(phen-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)NH(CH₂)₅—;-   —(CH₂)₃NHC(O)NH(CH₂)₅—;-   —(CH₂)₂C(O)NHCH₂(cyclohex-1,3-ylene)CH₂—;-   —(CH₂)₂NHC(O)(cyclopent-1,3-ylene)-;-   —(CH₂)₂NHC(O)NH(phen-1,4-ylene)(CH₂)₂—;-   1-[—(CH₂)₂C(O)](piperidin-4-yl)(CH₂)₂—;-   —(CH₂)₂NHC(O)(trans-cyclohex-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)-;-   —(CH₂)₂NH(phen-1,4-ylene)(CH₂)₂—;-   1-[—(CH₂)₂NHC(O)](piperidin-4-yl)(CH₂)₂—;-   —CH₂(phen-1,4-ylene)NH(phen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)CH₂—;-   —(CH₂)₂C(O)NHCH₂(pyrid-2,6-ylene)CH₂—;-   —(CH₂)₂C(O)NH(cis-cyclohex-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(trans-cyclohex-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)(phen-1,3-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)—((S)-isomer);-   —(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)—((R)-isomer);-   2-[(S)—(—CH₂—](pyrrolidin-1-yl)C(O)(CH₂)₄—;-   2-[(S)—(—CH₂—](pyrrolidin-1-yl)C(O)(phen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(4-chlorophen-1,3-ylene)CH₂—;-   —CH₂(2-fluorophen-1,3-ylene)CH₂—;-   —(CH₂)₂C(O)NH(4-methylphen-1,3-ylene)CH₂—;-   —(CH₂)₂C(O)NH(6-chlorophen-1,3-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2-chlorophen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2,6-dichlorophen-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)NHCH₂(phen-1,3-ylene)CH₂—;-   4-[—CH₂—](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)N(CH₂CH₃)(phen-1,4-ylene)CH₂—;-   1-[—(CH₂)₂NHC(O)](piperidin-4-yl)-;-   —(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂NHC(O)(thien-2,5-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)(3-nitrophen-1,4-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)-;-   1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)-;-   5-[—(CH₂)₂NHC(O)](pyrid-2-yl)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₃(thien-2,5-ylene)(CH₂)₃—;-   —(CH₂)₂(phen-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   —CH₂(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₂—;-   1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)CH₂—;-   —(CH₂)₂C(O)NH(3-chlorophen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2-(CF₃O—)phen-1,4-ylene)CH₂—;-   —(CH₂)₃(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂S(O)₂NH(CH₂)₅—;-   —CH₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂C(O)NH(2-iodophen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2-chloro-5-methoxyphen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2-chloro-6-methylphen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(CH₂)₅—;-   —(CH₂)₂N(CH₃)S(O)₂(phen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2-bromophen-1,4-ylene)CH₂—;-   —(CH₂)₃(phen-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₃(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₃—;-   —(CH₂)₂C(O)NH(2-methoxyphen-1,4-ylene)CH₂—;-   —(CH₂)₅NH(phen-1,4-ylene)(CH₂)₂—;-   4-[—(CH₂)₂—](piperidin-1-yl)(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂C(O)NH(phen-1,4-ylene)CH(CH₃)CH₂—;-   —(CH₂)₂-(trans-cyclohex-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂C(O)NH(2-fluorophen-1,4-ylene)CH₂—;-   —(CH₂)₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂C(O)NH(2,5-difluorophen-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)(phen-1,4-ylene)(CH₂)₂—;-   1-[—CH₂(pyrid-2,6-ylene)CH₂](piperidin-4-yl)CH₂—;-   —(CH₂)₃NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂NH(naphth-1,4-ylene)(CH₂)₂—;-   —(CH₂)₃O(phen-1,4-ylene)CH₂—;-   1-[—(CH₂)₃](piperidin-4-yl)CH₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂—;-   —(CH₂)₃(phen-1,4-ylene)NHC(O)(CH₂)₂—;-   —(CH₂)₃O(phen-1,4-ylene)(CH₂)₂—;-   2-[—(CH₂)₂](benzimidazol-5-yl)CH₂—;-   —(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₂—;-   —(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₄—;-   —(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₅—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₂—;-   —(CH₂)₂NHC(O)NH(phen-1,4-ylene)CH₂—;-   —(CH₂)₂N(CH₃)(CH₂)₂(cis-cyclohex-1,4-ylene)-;-   —(CH₂)₂C(O)NH(2,3,5,6-tetrafluorophen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2,6-diiodophen-1,4-ylene)CH₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₃—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₄—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₅—;-   —(CH₂)₂C(O)NHCH₂(phen-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)NHCH₂(phen-1,4-ylene)CH₂—;-   —(CH₂)₂C(O)NH(2-methylphen-1,4-ylene)CH₂—;-   1-[—(CH₂)₃O(phen-1,4-ylene)(CH₂)₂](piperidin-4-yl)CH₂—;-   —(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)(CH₂)₂—;-   —(CH₂)₂O(phen-1,3-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,4-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,3-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)(fur-2,5-ylene)CH₂—;-   —(CH₂)₂N(CH₃)C(O)(thien-2,5-ylene)CH₂—;-   —(CH₂)₂O(phen-1,4-ylene)O(CH₂)₂—;-   —(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,4-ylene)CH₂—;-   —(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,2-ylene)CH₂—;-   —(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,3-ylene)CH₂—;-   —(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,4-ylene)CH₂—;-   —(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂—;-   —(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(thien-2,5-ylene)CH₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,2-ylene)CH₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,3-ylene)CH₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,4-ylene)CH₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)(fur-2,5-ylene)CH₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)(thien-2,5-ylene)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,3-ylene)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,4-ylene)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,2-ylene)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,3-ylene)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,4-ylene)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂—;-   —(CH₂)₂(phen-1,4-ylene)NHC(O)(thien-2,5-ylene)CH₂—;-   —(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,3-ylene)CH₂—;-   —(CH₂)₃O(phen-1,3-ylene)CH₂—;-   —CH₂CH(OH)CH₂NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₄NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂NHC(O)CH₂—;-   —(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂NHC(O)CH₂—;-   —(CH₂)₂C(O)NHCH₂(trans-cyclohex-1,4-ylene)CH₂—;-   —(CH₂)₂NHC(O)(CH₂)₅—;-   —(CH₂)₂O(phen-1,3-ylene)O(CH₂)₂—;-   —(CH₂)₂O(phen-1,2-ylene)O(CH₂)₂—;-   —CH₂(phen-1,2-ylene)O(phen-1,2-ylene)CH₂—;-   —(CH₂)₂C(O)NH(CH₂)₆—;-   —(CH₂)₃(phen-1,4-ylene)(CH₂)₃—;-   —(CH₂)₃(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₄(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₃(furan-2,5-ylene)(CH₂)₃—;-   —(CH₂)₂N(CH₃)C(O)NH(phen-1,4-ylene)(CH₂)₂—;-   4-[—(CH₂)₂](piperidin-1-yl)C(O)NH(phen-1,4-ylene)(CH₂)₂—;-   —(CH₂)₃(phen-1,3-ylene)(CH₂)₃—;-   —(CH₂)₃(tetrahydrofuran-2,5-ylene)(CH₂)₃—; and-   —(CH₂)₂O(phen-1,4-ylene)C(O)(CH₂)₂—.    Representative Subgeneric Groupings

The following subgeneric formulae and groupings are intended to providerepresentative examples of various aspects and embodiments of thisinvention and as such, they are not intended to exclude otherembodiments or to limit the scope of this invention unless otherwiseindicated.

A particular group of compounds of formula I are those disclosed in U.S.Provisional Application No. 60/459,291, filed on Apr. 1, 2003. Thisgroup includes compounds of formula I; wherein:

Ar¹ represents a phenyl, (5-6C)cycloalkyl, (4-5C)heteroaryl or(4-5C)heterocyclyl group wherein the (4-5C)heteroaryl or(4-5C)heterocyclyl group contains one ring heteroatom selected fromoxygen, nitrogen and sulfur;

each R¹ represents an optional substituent on Ar¹ that is independentlyselected from the group consisting of (1-4C)alkyl, (2-4C)alkenyl,(2-4C)alkynyl, (3-6C)cycloalkyl, cyano, halo, —OR^(1a), —SR^(1b),—S(O)R^(1c), —S(O)₂R^(1d), and —NR^(1e)R^(1f); or two adjacent R¹ groupstogether form (3-6C)alkylene, (2-4C)alkylene-O— or—O-(1-4C)alkylene)-O—; wherein each alkyl, alkenyl or cycloalkyl groupis optionally substituted with from 1 to 5 fluorine atoms;

each of R^(1a), R^(1b), R^(1c), R^(1d), R^(1e) and R^(1f) isindependently hydrogen or (1-4C)alkyl;

a is 0 or an integer of from 1 to 3;

Ar² represents a phenyl, (5-6C)cycloalkyl, (4-5C)heteroaryl or(4-5C)heterocyclyl group wherein the (4-5C)heteroaryl or(4-5C)heterocyclyl group contains one ring heteroatom selected fromoxygen, nitrogen and sulfur;

each R² represents an optional substituent on Ar² that is independentlyselected from the group consisting of (1-4C)alkyl, (2-4C)alkenyl,(2-4C)alkynyl, (3-6C)cycloalkyl, cyano, halo, —OR^(2a), SR^(2b),—S(O)R^(2c), —S(O)₂R^(2d), and —NR^(2e)R^(2f); or two adjacent R² groupstogether form (3-6C)alkylene, (2-4C)alkylene-O— or—O-(1-4C)alkylene)-O—; wherein each alkyl, alkenyl or cycloalkyl groupis optionally substituted with from 1 to 5 fluorine atoms;

each of R^(2a), R^(2b), R^(2c), R^(2d), R^(2e) and R^(2f) isindependently hydrogen or (1-4C)alkyl;

b is 0 or an integer of from 1 to 3;

E is CN or C(O)NW^(a)W^(b);

each of W^(a) and W^(b) is independently selected from hydrogen and(1-4C)alkyl, or together with the nitrogen atom to which they areattached form a pyrrolidin-1-yl, piperidin-1-yl, piperazin-1-yl,morpholin-4-yl or thiomorpholin-4-yl group;

c is 0 or an integer of from 1 to 4;

each R³ is a substituent on carbon independently selected from the groupconsisting of (1-4C)alkyl and fluoro, wherein each alkyl group isoptionally substituted with from 1 to 5 fluorine atoms;

z is 1 or 2, the atom bearing the group E being attached to the ringcontaining the nitrogen atom at the 2- or 3-position with respect to thenitrogen atom;

R⁴ is a divalent group of the formula:—(R^(4a))_(d)-(A¹)_(e)-(R^(4b))_(f)-Q-(R^(4c))_(g)-(A²)_(h)-(R^(4d))_(i)—wherein

d, e, f, g, h and i are each independently selected from 0 and 1;

R^(4a), R^(4b), R^(4c) and R^(4d) are each independently selected fromthe group consisting of (1-10C)alkylene, (2-10C)alkenylene and(2-10C)alkynylene wherein each alkylene, alkenylene or alkynylene groupis unsubstituted or substituted with from 1 to 5 substituentsindependently selected from the group consisting of (1-4C)alkyl, fluoro,hydroxy, phenyl and phenyl(1-4C)-alkyl;

A¹ and A² are each independently selected from (3-7C)cycloalkylene,(6-10C)arylene, (2-9C)heteroarylene and (3-6C)heterocyclene; whereineach cycloalkylene is unsubstituted or substituted with from 1 to 4substitutents selected independently from (1-4C)alkyl and each arylene,heteroarylene or heterocyclene group is unsubstituted or substitutedwith from 1 to 4 substituents independently selected from the groupconsisting of halogen, (1-4C)alkyl and (1-4C)alkoxy;

Q is selected from the group consisting of a bond, —O—, —C(O)O—,—OC(O)—, —S—, —S(O)—, —S(O)₂—, —N(Q^(a))C(O)—, —C(O)N(Q^(b))-,—N(Q^(c))S(O)₂—, —S(O)₂N(Q^(d))-, —N(Q^(e))C(O)N(Q^(f))-,—N(Q^(g))S(O)₂N(Q^(h))-, —OC(O)N(Q^(i))- and —N(Q^(j))C(O)O—;

Q^(a), Q^(b), Q^(c), Q^(d), Q^(e), Q^(f), Q^(g), Q^(h), Q^(i) and Q^(j)are each independently selected from the group consisting of hydrogen,(1-6C)alkyl, A³ and (1-4C)alkylene-A⁴; wherein the alkyl group isunsubstituted or substituted with from 1 to 3 substituents independentlyselected from fluoro, hydroxy and (1-4C)alkoxy; or together with thenitrogen atom and the group R^(4b) or R^(4c) to which they are attached,form a 4-6 membered azacycloalkylene group;

A³ and A⁴ are each independently selected from (3-6C)cycloalkyl,(6-10C)aryl, (2-9C)heteroaryl and (3-6C)heterocyclyl; wherein eachcycloalkyl is unsubstituted or substituted with from 1 to 4substitutents selected independently from (1-4C)alkyl and each aryl,heteroaryl or heterocyclyl group is unsubstituted or substituted withfrom 1 to 4 substituents independently selected from the groupconsisting of halogen, (1-4C)alkyl and (1-4C)alkoxy;

provided that the number of contiguous atoms in the shortest chainbetween the two nitrogen atoms to which R⁴ is attached is in the rangeof from 4 to 14;

R⁵ represents hydrogen or (1-4C)alkyl;

R⁶ is —NR^(6a)CR^(6b)(O) or —CR^(6c)R^(6d)OR^(6e) and R⁷ is hydrogen, orR⁶ and R⁷ together form —NR^(7a)C(O)—CR^(7b)═CR^(7c)—,—CR^(7d)═CR^(7e)C(O)—NR^(7f)—, —NR^(7g)C(O)—CR^(7h)R^(7i)—CR^(7j)R^(7k)—or —CR^(7l)R^(7m)—CR^(7n)R^(7o)—C(O)—NR^(7p)—;

each of R^(6a), R^(6b), R^(6c), R^(6d) and R^(6e) is independentlyhydrogen or (1-4C)alkyl; and

each of R^(7a), R^(7b), R^(7c), R^(7d), R^(7e), R^(7f), R^(7g), R^(7h),R^(7i), R^(7j), R^(7k), R^(7l), R^(7m), R^(7n), R^(7o) and R^(7p) isindependently hydrogen or (1-4C)alkyl;

or a pharmaceutically-acceptable salt or solvate or stereoisomerthereof.

Another particular group of compounds of formula I are those where: a is0; b is 0; c is 0; R⁵ is hydrogen; and E, Ar¹, Ar², R⁴, R⁶, R⁷ and z areas defined herein; or a pharmaceutically acceptable salt or solvate orstereoisomer thereof.

Yet another particular group of compounds of formula I are those where:c is 0; z is 1; R⁵ is hydrogen; E is —C(O)NW^(a)W^(b); Ar¹ is phenyl;Ar² is phenyl; and R¹, R², R⁴, R⁶, R⁷, W^(a), W^(b), a and b are asdefined herein; or a pharmaceutically acceptable salt or solvate orstereoisomer thereof.

Still another particular group of compounds of formula I are thosewhere: c is 0; z is 1; R⁵ is hydrogen; E is —C(O)NH₂; Ar¹ is phenyl; Ar²is phenyl; and R¹, R², R⁴, R⁶, R⁷, a and b are as defined herein; or apharmaceutically acceptable salt or solvate or stereoisomer thereof.

Another particular group of compounds of formula I are those of formulaII as defined herein; or a pharmaceutically acceptable salt or solvateor stereoisomer thereof.

Another particular group of compounds of formula I are those of formulaIII as defined herein; or a pharmaceutically acceptable salt or solvateor stereoisomer thereof.

Another particular group of compounds of formula I are those of formulaIV as defined herein; or a pharmaceutically acceptable salt or solvateor stereoisomer thereof.

Another particular group of compounds of formula I are those of formulaV as defined herein; or a pharmaceutically acceptable salt or solvate orstereoisomer thereof.

Another particular group of compounds of formula I are compounds offormula IIIa:

wherein R⁴ is as defined in Table 1; or a pharmaceutically acceptablesalt or solvate thereof.

TABLE I Ex. R⁴ 1 —(CH₂)₉— (racemic)¹ 2 —(CH₂)₉— 3 —(CH₂)₂C(O)NH(CH₂)₅— 4—(CH₂)₂N(CH₃)C(O)(CH₂)₅— 5 —(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 6—(CH₂)₂NHC(O)(phen-1,4-ylene)CH₂— 7 —(CH₂)₂NHC(O)NH(CH₂)₅— 8—(CH₂)₃NHC(O)NH(CH₂)₅— 9 —(CH₂)₂C(O)NHCH₂(cyclohex-1,3-ylene)CH₂— 10—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)- 11—(CH₂)₂C(O)NH(2-chlorophen-1,4-ylene)CH₂— 12 —(CH₂)₂S(O)₂NH(CH₂)₅— 13—(CH₂)₂N(CH₃)S(O)₂(phen-1,4-ylene)CH₂— 14—(CH₂)₂NHC(O)NHCH₂(phen-1,3-ylene)CH₂— 15—(CH₂)₃(phen-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂— 161-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)CH₂— 17—(CH₂)₃O(phen-1,4-ylene)(CH₂)₂— 18 —(CH₂)₄(phen-1,4-ylene)(CH₂)₂— 19—(CH₂)₃(thien-2,5-ylene)(CH₂)₃— 20—(CH₂)₂C(O)NH(2-chloro-5-methoxyphen-1,4-ylene)CH₂— 21 —(CH₂)₇— 22—(CH₂)₈— 23 —(CH₂)₂NHC(O)NH(phen-1,4-ylene)(CH₂)₂— 241-[-(CH₂)₂C(O)](piperidin-4-yl)(CH₂)₂— 25—(CH₂)₂NHC(O)(trans-cyclohex-1,4-ylene)CH₂— 26—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)- 27—(CH₂)₂NH(phen-1,4-ylene)(CH₂)₂— 281-[-(CH₂)₂NHC(O)](piperidin-4-yl)(CH₂)₂— 29—CH₂(phen-1,4-ylene)NH(phen-1,4-ylene)CH₂— 30—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)CH₂— 31—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 32—(CH₂)₂C(O)NHCH₂(pyrid-2,6-ylene)CH₂— 33—(CH₂)₂C(O)NH(cis-cyclohex-1,4-ylene)CH₂— 34—(CH₂)₂C(O)NH(trans-cyclohex-1,4-ylene)CH₂— 35—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)CH₂— 36—(CH₂)₂N(CH₃)C(O)(phen-1,3-ylene)CH₂— 37—(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)CH₂— 38—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 39—(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)— ((S)-isomer) 40—(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)— ((R)-isomer) 41 2-[(S)

CH₂—](pyrrolidin-1-yl)C(O)(CH₂)₄— 42 2-[(S)

CH₂—](pyrrolidin-1-yl)C(O)(phen-1,4-ylene)CH₂— 43—(CH₂)₂C(O)NH(4-chlorophen-1,3-ylene)CH₂— 44—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 451-[-(CH₂)₂C(O)](piperidin-4-yl)(CH₂)₂— 46—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)CH₂— 47 —CH₂(2-fluorophen-1,3-ylene)CH₂—48 —(CH₂)₂C(O)NH(4-methylphen-1,3-ylene)CH₂— 49—(CH₂)₂C(O)NH(6-chlorophen-1,3-ylene)CH₂— 50—(CH₂)₂C(O)NH(2,6-dichlorophen-1,4-ylene)CH₂— 514-[-CH₂—](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂— 52—(CH₂)₂C(O)N(CH₂CH₃)(phen-1,4-ylene)CH₂— 531-[-(CH₂)₂NHC(O)](piperidin-4-yl)- 54—(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂— 55—(CH₂)₂NHC(O)(thien-2,5-ylene)CH₂— 56—(CH₂)₂N(CH₃)C(O)(3-nitrophen-1,4-ylene)CH₂— 57—(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)- 581-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)- 595-[-(CH₂)₂NHC(O)](pyrid-2-yl)CH₂— 60 1-[-(CH₂)₃](piperidin-4-yl)CH₂— 61—CH₂(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂— 621-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₂— 63—(CH₂)₃NH(phen-1,4-ylene)(CH₂)₂— 64—(CH₂)₂C(O)NH(3-chlorophen-1,4-ylene)CH₂— 65—(CH₂)₂C(O)NH(2-(CF₃O—)phen-1,4-ylene)CH₂— 66—(CH₂)₃(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂— 67—CH₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂— 68—(CH₂)₂C(O)NH(2-iodophen-1,4-ylene)CH₂— 69—(CH₂)₂C(O)NH(2-chloro-6-methylphen-1,4-ylene)CH₂— 70—(CH₂)₃C(O)NH(CH₂)₅— 71 —(CH₂)₂C(O)NH(2-bromophen-1,4-ylene)CH₂— 72—(CH₂)₃(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂— 731-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₃— 74—(CH₂)₂C(O)NH(2-methoxyphen-1,4-ylene)CH₂— 75—(CH₂)₅NH(phen-1,4-ylene)(CH₂)₂— 764-[-(CH₂)₂—](piperidin-1-yl)(phen-1,4-ylene)(CH₂)₂— 77—(CH₂)₂C(O)NH(phen-1,4-ylene)CH(CH₃)CH₂— 78—(CH₂)₂-(trans-cyclohex-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂— 79—(CH₂)₂C(O)NH(2-fluorophen-1,4-ylene)CH₂— 80—(CH₂)₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂— 81—(CH₂)₂C(O)NH(2,5-difluorophen-1,4-ylene)CH₂— 82—(CH₂)₂NHC(O)(phen-1,4-ylene)(CH₂)₂— 831-[-CH₂(pyrid-2,6-ylene)CH₂](piperidin-4-yl)CH₂— 84—(CH₂)₂NH(naphth-1,4-ylene)(CH₂)₂— 854-[-(CH₂)₂](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂— 86—(CH₂)₃(phen-1,4-ylene)NHC(O)(CH₂)₂— 87 —(CH₂)₃O(phen-1,4-ylene)CH₂— 882-[-(CH₂)₂](benzimidazol-5-yl)CH₂— 89—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₂— 90—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₄— 91—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₅— 924-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₂— 93—(CH₂)₂NHC(O)NH(phen-1,4-ylene)CH₂— 94—(CH₂)₂N(CH₃)(CH₂)₂(cis-cyclohex-1,4-ylene)- 95—(CH₂)₂C(O)NH(2,3,5,6-tetrafluorophen-1,4-ylene)CH₂— 96—(CH₂)₂C(O)NH(2,6-diiodophen-1,4-ylene)CH₂— 974-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₃— 984-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₄— 994-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₅— 100—(CH₂)₂C(O)NHCH₂(phen-1,4-ylene)CH₂— 101—(CH₂)₂NHC(O)NHCH₂(phen-1,4-ylene)CH₂— 192—(CH₂)₂C(O)NH(2-methylphen-1,4-ylene)CH₂— 1031-[-(CH₂)₃O(phen-1,4-ylene)(CH₂)₂](piperidin-4-yl)CH₂— 104—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)(CH₂)₂— 105 —(CH₂)₂O(phen-1,3-ylene)CH₂—106 —(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,4-ylene)CH₂— 107—(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,3-ylene)CH₂— 108—(CH₂)₂N(CH₃)C(O)(fur-2,5-ylene)CH₂— 109—(CH₂)₂N(CH₃)C(O)(thien-2,5-ylene)CH₂— 110—(CH₂)₂O(phen-1,4-ylene)O(CH₂)₂— 111—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,4- ylene)CH₂— 112—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,2- ylene)CH₂— 113—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,3- ylene)CH₂— 114—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,4- ylene)CH₂— 115—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂— 116—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(thien-2,5- ylene)CH₂— 1174-[-(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,2-ylene)CH₂— 1184-[-(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,3-ylene)CH₂— 1194-[-(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,4-ylene)CH₂— 1204-[-(CH₂)₂](piperidin-1-yl)C(O)(fur-2,5-ylene)CH₂— 1214-[-(CH₂)₂](piperidin-1-yl)C(O)(thien-2,5-ylene)CH₂— 122—(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,3-ylene)CH₂— 123—(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,4-ylene)CH₂— 124—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,2-ylene)CH₂— 125—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,3-ylene)CH₂— 126—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,4-ylene)CH₂— 127—(CH₂)₂(phen-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂— 128—(CH₂)₂(phen-1,4-ylene)NHC(O)(thien-2,5-ylene)CH₂— 129—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,3- ylene)CH₂— 130—(CH₂)₃O(phen-1,3-ylene)CH₂— 131 —CH₂CH(OH)CH₂NH(phen-1,4-ylene)(CH₂)₂—132 —(CH₂)₄NH(phen-1,4-ylene)(CH₂)₂— 133—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂NHC(O)CH₂— 134—(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂NHC(O)CH₂— 135—(CH₂)₂C(O)NHCH₂(trans-cyclohex-1,4-ylene)CH₂— 136 —(CH₂)₂NHC(O)(CH₂)₅—137 —(CH₂)₂O(phen-1,3-ylene)O(CH₂)₂— 138—(CH₂)₂O(phen-1,2-ylene)O(CH₂)₂— 139—CH₂(phen-1,2-ylene)O(phen-1,2-ylene)CH₂— 140 —(CH₂)₂C(O)NH(CH₂)₆— 141—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)- 142—(CH₂)₃(phen-1,4-ylene)(CH₂)₃— 143 —(CH₂)₃(phen-1,4-ylene)(CH₂)₂— 144—(CH₂)₄(phen-1,4-ylene)(CH₂)₂— 145 —(CH₂)₃(furan-2,5-ylene)(CH₂)₃— 146—(CH₂)₂N(CH₃)C(O)NH(phen-1,4-ylene)(CH₂)₂— 1474-[-(CH₂)₂](piperidin-1-yl)C(O)NH(phen-1,4-ylene)(CH₂)₂— 148—(CH₂)₃(phen-1,3-ylene)(CH₂)₃— 149—(CH₂)₃(tetrahydrofuran-2,5-ylene)(CH₂)₃— 150—(CH₂)₂O(phen-1,4-ylene)C(O)(CH₂)₂— ¹This compound is racemic at thechiral carbon bearing the hydroxyl group in formula IIIa.

Another particular group of compounds of formula I are compounds offormula IVa:

wherein R⁴ is as defined in Table II; or a pharmaceutically acceptablesalt or solvate thereof.

TABLE II Ex. R⁴ 151 —(CH₂)₉— 152 —(CH₂)₁₀— 153 —(CH₂)₂C(O)NH(CH₂)₅— 154—(CH₂)₂N(CH₃)C(O)(CH₂)₅— 155 —(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 156—(CH₂)₂NHC(O)(phen-1,4-ylene)CH₂— 157 —(CH₂)₂NHC(O)NH(CH₂)₅— 158—(CH₂)₃NHC(O)NH(CH₂)₅— 159 —(CH₂)₂C(O)NHCH₂(cyclohex-1,3-ylene)CH₂— 160—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)- 161—(CH₂)₂C(O)NH(2-chlorophen-1,4-ylene)CH₂— 162 —(CH₂)₂S(O)₂NH(CH₂)₅— 163—(CH₂)₂N(CH₃)S(O)₂(phen-1,4-ylene)CH₂— 164—(CH₂)₂NHC(O)NHCH₂(phen-1,3-ylene)CH₂— 165—(CH₂)₃(phen-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂— 1661-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)CH₂— 167—(CH₂)₃O(phen-1,4-ylene)(CH₂)₂— 168 —(CH₂)₄(phen-1,4-ylene)(CH₂)₂— 169—(CH₂)₃(thien-2,5-ylene)(CH₂)₃— 170—(CH₂)₂C(O)NH(2-chloro-5-methoxyphen-1,4-ylene)CH₂— 171 —(CH₂)₇— 172—(CH₂)₈— 173 —(CH₂)₂NHC(O)NH(phen-1,4-ylene)(CH₂)₂— 1741-[-(CH₂)₂C(O)](piperidin-4-yl)(CH₂)₂— 175—(CH₂)₂NHC(O)(trans-cyclohex-1,4-ylene)CH₂— 176—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)- 177—(CH₂)₂NH(phen-1,4-ylene)(CH₂)₂— 1781-[-(CH₂)₂NHC(O)](piperidin-4-yl)(CH₂)₂— 179—CH₂(phen-1,4-ylene)NH(phen-1,4-ylene)CH₂— 180—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)CH₂— 181—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 182—(CH₂)₂C(O)NHCH₂(pyrid-2,6-ylene)CH₂— 183—(CH₂)₂C(O)NH(cis-cyclohex-1,4-ylene)CH₂— 184—(CH₂)₂C(O)NH(trans-cyclohex-1,4-ylene)CH₂— 185—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)CH₂— 186—(CH₂)₂N(CH₃)C(O)(phen-1,3-ylene)CH₂— 187—(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)CH₂— 188—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 189—(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)— ((S)-isomer) 190—(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)— ((R)-isomer) 191 2-[(S)

CH₂—](pyrrolidin-1-yl)C(O)(CH₂)₄— 192 2-[(S)

CH₂—](pyrrolidin-1-yl)C(O)(phen-1,4-ylene)CH₂— 193—(CH₂)₂C(O)NH(4-chlorophen-1,3-ylene)CH₂— 194—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂— 1951-[-(CH₂)₂C(O)](piperidin-4-yl)(CH₂)₂— 196—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)CH₂— 197—CH₂(2-fluorophen-1,3-ylene)CH₂— 198—(CH₂)₂C(O)NH(4-methylphen-1,3-ylene)CH₂— 199—(CH₂)₂C(O)NH(6-chlorophen-1,3-ylene)CH₂— 200—(CH₂)₂C(O)NH(2,6-dichlorophen-1,4-ylene)CH₂— 2014-[-CH₂—](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂— 202—(CH₂)₂C(O)N(CH₂CH₃)(phen-1,4-ylene)CH₂— 2031-[-(CH₂)₂NHC(O)](piperidin-4-yl)- 204—(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂— 205—(CH₂)₂NHC(O)(thien-2,5-ylene)CH₂— 206—(CH₂)₂N(CH₃)C(O)(3-nitrophen-1,4-ylene)CH₂— 207—(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)- 2081-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)- 2095-[-(CH₂)₂NHC(O)](pyrid-2-yl)CH₂— 210 1-[-(CH₂)₃](piperidin-4-yl)CH₂—211 —CH₂(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂— 2121-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₂— 213—(CH₂)₃NH(phen-1,4-ylene)(CH₂)₂— 214—(CH₂)₂C(O)NH(3-chlorophen-1,4-ylene)CH₂— 215—(CH₂)₂C(O)NH(2-(CF₃O-)phen-1,4-ylene)CH₂— 216—(CH₂)₃(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂— 217—CH₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂— 218—(CH₂)₂C(O)NH(2-iodophen-1,4-ylene)CH₂— 219—(CH₂)₂C(O)NH(2-chloro-6-methylphen-1,4-ylene)CH₂— 220—(CH₂)₃C(O)NH(CH₂)₅— 221 —(CH₂)₂C(O)NH(2-bromophen-1,4-ylene)CH₂— 222—(CH₂)₃(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂— 2231-[-CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₃— 224—(CH₂)₂C(O)NH(2-methoxyphen-1,4-ylene)CH₂— 225—(CH₂)₅NH(phen-1,4-ylene)(CH₂)₂— 2264-[-(CH₂)₂—](piperidin-1-yl)(phen-1,4-ylene)(CH₂)₂— 227—(CH₂)₂C(O)NH(phen-1,4-ylene)CH(CH₃)CH₂— 228—(CH₂)₂-(trans-cyclohex-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂— 229—(CH₂)₂C(O)NH(2-fluorophen-1,4-ylene)CH₂— 230—(CH₂)₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂— 231—(CH₂)₂C(O)NH(2,5-difluorophen-1,4-ylene)CH₂— 232—(CH₂)₂NHC(O)(phen-1,4-ylene)(CH₂)₂— 2331-[-CH₂(pyrid-2,6-ylene)CH₂](piperidin-4-yl)CH₂— 234—(CH₂)₂NH(naphth-1,4-ylene)(CH₂)₂— 2354-[-(CH₂)₂](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂— 236—(CH₂)₃(phen-1,4-ylene)NHC(O)(CH₂)₂— 237 —(CH₂)₃O(phen-1,4-ylene)CH₂—238 2-[-(CH₂)₂](benzimidazol-5-yl)CH₂— 239—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₂— 240—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₄— 241—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₅— 2424-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₂— 243—(CH₂)₂NHC(O)NH(phen-1,4-ylene)CH₂— 244—(CH₂)₂N(CH₃)(CH₂)₂(cis-cyclohex-1,4-ylene)- 245—(CH₂)₂C(O)NH(2,3,5,6-tetrafluorophen-1,4-ylene)CH₂— 246—(CH₂)₂C(O)NH(2,6-diiodophen-1,4-ylene)CH₂— 2474-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₃— 2484-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₄— 2494-[-(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₅— 250—(CH₂)₂C(O)NHCH₂(phen-1,4-ylene)CH₂— 251—(CH₂)₂NHC(O)NHCH₂(phen-1,4-ylene)CH₂— 252—(CH₂)₂C(O)NH(2-methylphen-1,4-ylene)CH₂— 2531-[-(CH₂)₃O(phen-1,4-ylene)(CH₂)₂](piperidin-4-yl)CH₂— 254—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)(CH₂)₂— 255 —(CH₂)₂O(phen-1,3-ylene)CH₂—256 —(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,4-ylene)CH₂— 257—(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,3-ylene)CH₂— 258—(CH₂)₂N(CH₃)C(O)(fur-2,5-ylene)CH₂— 259—(CH₂)₂N(CH₃)C(O)(thien-2,5-ylene)CH₂— 260—(CH₂)₂O(phen-1,4-ylene)O(CH₂)₂— 261—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,4- ylene)CH₂— 262—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,2- ylene)CH₂— 263—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,3- ylene)CH₂— 264—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,4- ylene)CH₂— 265—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂— 266—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(thien-2,5- ylene)CH₂— 2674-[-(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,2-ylene)CH₂— 2684-[-(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,3-ylene)CH₂— 2694-[-(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,4-ylene)CH₂— 2704-[-(CH₂)₂](piperidin-1-yl)C(O)(fur-2,5-ylene)CH₂— 2714-[-(CH₂)₂](piperidin-1-yl)C(O)(thien-2,5-ylene)CH₂— 272—(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,3-ylene)CH₂— 273—(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,4-ylene)CH₂— 274—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,2-ylene)CH₂— 275—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,3-ylene)CH₂— 276—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,4-ylene)CH₂— 277—(CH₂)₂(phen-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂— 278—(CH₂)₂(phen-1,4-ylene)NHC(O)(thien-2,5-ylene)CH₂— 279—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,3- ylene)CH₂— 280—(CH₂)₃O(phen-1,3-ylene)CH₂— 281 —CH₂CH(OH)CH₂NH(phen-1,4-ylene)(CH₂)₂—282 —(CH₂)₄NH(phen-1,4-ylene)(CH₂)₂— 283—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂NHC(O)CH₂— 284—(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂NHC(O)CH₂— 285—(CH₂)₂C(O)NHCH₂(trans-cyclohex-1,4-ylene)CH₂— 286 —(CH₂)₂NHC(O)(CH₂)₅—287 —(CH₂)₂O(phen-1,3-ylene)O(CH₂)₂— 288—(CH₂)₂O(phen-1,2-ylene)O(CH₂)₂— 289—CH₂(phen-1,2-ylene)O(phen-1,2-ylene)CH₂— 290 —(CH₂)₂C(O)NH(CH₂)₆— 291—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)- 292—(CH₂)₃(phen-1,4-ylene)(CH₂)₃— 293 —(CH₂)₃(phen-1,4-ylene)(CH₂)₂— 294—(CH₂)₄(phen-1,4-ylene)(CH₂)₂— 295 —(CH₂)₃(furan-2,5-ylene)(CH₂)₃— 296—(CH₂)₂N(CH₃)C(O)NH(phen-1,4-ylene)(CH₂)₂— 2974-[-(CH₂)₂](piperidin-1-yl)C(O)NH(phen-1,4-ylene)(CH₂)₂— 298—(CH₂)₃(phen-1,3-ylene)(CH₂)₃— 299—(CH₂)₃(tetrahydrofuran-2,5-ylene)(CH₂)₃— 300—(CH₂)₂O(phen-1,4-ylene)C(O)(CH₂)₂—Definitions

When describing the compounds, compositions, methods and processes ofthis invention, the following terms have the following meanings unlessotherwise indicated.

The term “alkyl” means a monovalent saturated hydrocarbon group whichmay be linear or branched. Unless otherwise defined, such alkyl groupstypically contain from 1 to 10 carbon atoms. Representative alkyl groupsinclude, by way of example, methyl, ethyl, n-propyl, isopropyl, n-butyl,sec-butyl, isobutyl, tert-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl,n-nonyl, n-decyl and the like.

The term “alkylene” means a divalent saturated hydrocarbon group whichmay be linear or branched. Unless otherwise defined, such alkylenegroups typically contain from 1 to 10 carbon atoms. Representativealkylene groups include, by way of example, methylene, ethane-1,2-diyl(“ethylene”), propane-1,2-diyl, propane-1,3-diyl, butane-1,4-diyl,pentane-1,5-diyl and the like.

The term “alkoxy” means a monovalent group of the formula (alkyl)-O—,where alkyl is as defined herein. Representative alkoxy groups include,by way of example, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy,sec-butoxy, isobutoxy, tert-butoxy and the like.

The term “alkenyl” means a monovalent unsaturated hydrocarbon groupwhich may be linear or branched and which has at least one, andtypically 1, 2 or 3, carbon-carbon double bonds. Unless otherwisedefined, such alkenyl groups typically contain from 2 to 10 carbonatoms. Representative alkenyl groups include, by way of example,ethenyl, n-propenyl, isopropenyl, n-but-2-enyl, n-hex-3-enyl and thelike. The term “alkenylene” means a divalent alkenyl group.

The term “alkynyl” means a monovalent unsaturated hydrocarbon groupwhich may be linear or branched and which has at least one, andtypically 1, 2 or 3, carbon-carbon triple bonds. Unless otherwisedefined, such alkynyl groups typically contain from 2 to 10 carbonatoms. Representative alkynyl groups include, by way of example,ethynyl, n-propynyl, n-but-2-ynyl, n-hex-3-ynyl and the like. The term“alkynylene” means a divalent alkynyl group.

The term “aryl” means a monovalent aromatic hydrocarbon having a singlering (i.e., phenyl) or fused rings (i.e., naphthalene). Unless otherwisedefined, such aryl groups typically contain from 6 to 10 carbon ringatoms. Representative aryl groups include, by way of example, phenyl andnaphthalene-1-yl, naphthalene-2-yl, and the like. The term “arylene”means a divalent aryl group.

The term “azacycloalkyl” means a monovalent heterocyclic ring containingone nitrogen atom, i.e., a cycloalkyl group in which one carbon atom hasbeen replaced with a nitrogen atom. Unless otherwise defined, suchazacycloalkyl groups typically contain from 2 to 9 carbon atoms.Representative examples of an azacycloalkyl group are pyrrolidinyl andpiperidinyl groups. The term “azacycloalkylene” means a divalentazacycloakyl group. Representative examples of an azacycloalkylene groupare pyrrolidinylene and piperidinylene groups.

The term “cycloalkyl” means a monovalent saturated carbocyclichydrocarbon group. Unless otherwise defined, such cycloalkyl groupstypically contain from 3 to 10 carbon atoms. Representative cycloalkylgroups include, by way of example, cyclopropyl, cyclobutyl, cyclopentyl,cyclohexyl and the like. The term “cycloalkylene” means a divalentcycloalkyl group.

The term “halo” means fluoro, chloro, bromo and iodo.

The term “heteroaryl” means a monovalent aromatic group having a singlering or two fused rings and containing in the ring at least oneheteroatom (typically 1 to 3 heteroatoms) selected from nitrogen, oxygenor sulfur. Unless otherwise defined, such heteroaryl groups typicallycontain from 5 to 10 total ring atoms. Representative heteroaryl groupsinclude, by way of example, monovalent species of pyrrole, imidazole,thiazole, oxazole, furan, thiophene, triazole, pyrazole, isoxazole,isothiazole, pyridine, pyrazine, pyridazine, pyrimidine, triazine,indole, benzofuran, benzothiophene, benzimidazole, benzthiazole,quinoline, isoquinoline, quinazoline, quinoxaline and the like, wherethe point of attachment is at any available carbon or nitrogen ringatom. The term “heteroarylene” means a divalent heteroaryl group.

The term “heterocyclyl” or “heterocyclic” means a monovalent saturatedor unsaturated (non-aromatic) group having a single ring or multiplecondensed rings and containing in the ring at least one heteroatom(typically 1 to 3 heteroatoms) selected from nitrogen, oxygen or sulfur.Unless otherwise defined, such heterocyclic groups typically containfrom 2 to 9 total ring carbon atoms. Representative heterocyclic groupsinclude, by way of example, monovalent species of pyrrolidine,imidazolidine, pyrazolidine, piperidine, 1,4-dioxane, morpholine,thiomorpholine, piperazine, 3-pyrroline and the like, where the point ofattachment is at any available carbon or nitrogen ring atom. The term“heterocyclene” means a divalent heterocyclyl or heterocyclic group.

When a specific number of carbon atoms is intended for a particular termused herein, the number of carbon atoms is shown in parenthesespreceding the term. For example, the term “(1-4C)alkyl” means an alkylgroup having from 1 to 4 carbon atoms.

The term “pharmaceutically acceptable salt” means a salt which isacceptable for administration to a patient, such as a mammal (e.g.,salts having acceptable mammalian safety for a given dosage regime).Such salts can be derived from pharmaceutically acceptable inorganic ororganic bases and from pharmaceutically acceptable inorganic or organicacids. Salts derived from pharmaceutically acceptable inorganic basesinclude ammonium, calcium, copper, ferric, ferrous, lithium, magnesium,manganic, manganous, potassium, sodium, zinc and the like. Particularlypreferred are ammonium, calcium, magnesium, potassium and sodium salts.Salts derived from pharmaceutically acceptable organic bases includesalts of primary, secondary and tertiary amines, including substitutedamines, cyclic amines, naturally-occurring amines and the like, such asarginine, betaine, caffeine, choline, N,N′-dibenzylethylenediamine,diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol,ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine,glucamine, glucosamine, histidine, hydrabamine, isopropylamine, lysine,methylglucamine, morpholine, piperazine, piperadine, polyamine resins,procaine, purines, theobromine, triethylamine, trimethylamine,tripropylamine, tromethamine and the like. Salts derived frompharmaceutically acceptable acids include acetic, ascorbic,benzenesulfonic, benzoic, camphosulfonic, citric, ethanesulfonic,edisylic, fumaric, gentisic, gluconic, glucoronic, glutamic, hippuric,hydrobromic, hydrochloric, isethionic, lactic, lactobionic, maleic,malic, mandelic, methanesulfonic, mucic, naphthalenesulfonic,naphthalene-1,5-disulfonic, naphthalene-2,6-disulfonic, nicotinic,nitric, orotic, pamoic, pantothenic, phosphoric, succinic, sulfuric,tartaric, p-toluenesulfonic, xinafoic and the like. Particularlypreferred are citric, hydrobromic, hydrochloric, isethionic, maleic,naphthalene-1,5-disulfonic, phosphoric, sulfuric and tartaric acids.

The term “salt thereof” means a compound formed when the hydrogen of anacid is replaced by a cation, such as a metal cation or an organiccation and the like. Preferably, the salt is a pharmaceuticallyacceptable salt, although this is not required for salts of intermediatecompounds that are not intended for administration to a patient.

The term “solvate” means a complex or aggregate formed by one or moremolecules of a solute, i.e. a compound of formula I or apharmaceutically acceptable salt thereof, and one or more molecules of asolvent. Such solvates are typically crystalline solids having asubstantially fixed molar ratio of solute and solvent. Representativesolvents include, by way of example, water, methanol, ethanol,isopropanol, acetic acid and the like. When the solvent is water, thesolvate formed is a hydrate.

It will be appreciated that the term “or a pharmaceutically acceptablesalt or solvate or stereoisomer thereof” is intended to include allpermutations of salts, solvates and stereoisomers, such as a solvate ofa pharmaceutically acceptable salt of a stereoisomer of a compound offormula I.

The term “therapeutically effective amount” means an amount sufficientto effect treatment when administered to a patient in need of treatment.

The term “treating” or “treatment” as used herein means the treating ortreatment of a disease or medical condition (such as COPD) in a patient,such as a mammal (particularly a human) that includes:

-   -   (a) preventing the disease or medical condition from occurring,        i.e., prophylactic treatment of a patient;    -   (b) ameliorating the disease or medical condition, i.e.,        eliminating or causing regression of the disease or medical        condition in a patient;    -   (c) suppressing the disease or medical condition, i.e., slowing        or arresting the development of the disease or medical condition        in a patient; or    -   (d) alleviating the symptoms of the disease or medical condition        in a patient.

The term “leaving group” means a functional group or atom which can bedisplaced by another functional group or atom in a substitutionreaction, such as a nucleophilic substitution reaction. By way ofexample, representative leaving groups include chloro, bromo and iodogroups; sulfonic ester groups, such as mesylate, tosylate, brosylate,nosylate and the like; and acyloxy groups, such as acetoxy,trifluoroacetoxy and the like.

The term “protected derivatives thereof” means a derivative of thespecified compound in which one or more functional groups of thecompound are protected from undesired reactions with a protecting orblocking group. Functional groups which may be protected include, by wayof example, carboxylic acid groups, amino groups, hydroxyl groups, thiolgroups, carbonyl groups and the like. Representative protecting groupsfor carboxylic acids include esters (such as p-methoxybenzyl ester),amides and hydrazides; for amino groups, carbamates (such astert-butoxycarbonyl) and amides; for hydroxyl groups, ethers and esters;for thiol groups, thioethers and thioesters; for carbonyl groups,acetals and ketals; and the like. Such protecting groups are well-knownto those skilled in the art and are described, for example, in T. W.Greene and G. M. Wuts, Protecting Groups in Organic Synthesis, ThirdEdition, Wiley, New York, 1999, and references cited therein.

The term “amino-protecting group” means a protecting group suitable forpreventing undesired reactions at an amino group. Representativeamino-protecting groups include, but are not limited to,tert-butoxycarbonyl (BOC), trityl (Tr), benzyloxycarbonyl (Cbz),9-fluorenylmethoxycarbonyl (Fmoc), formyl, trimethylsilyl (TMS),tert-butyldimethylsilyl (TBS), and the like.

The term “carboxy-protecting group” means a protecting group suitablefor preventing undesired reactions at a carboxy group. Representativecarboxy-protecting groups include, but are not limited to, esters, suchas methyl, ethyl, tert-butyl, benzyl (Bn), p-methoxybenzyl (PMB),9-fluoroenylmethyl (Fm), trimethylsilyl (TMS), tert-butyldimethylsilyl(TBS), diphenylmethyl (benzhydryl, DPM) and the like.

The term “hydroxyl-protecting group” means a protecting group suitablefor preventing undesirable reactions at a hydroxyl group. Representativehydroxyl-protecting groups include, but are not limited to, silyl groupsincluding tri(1-6C)alkylsilyl groups, such as trimethylsilyl (TMS),triethylsilyl (TES), tert-butyldimethylsilyl (TBS) and the like; esters(acyl groups) including (1-6C)alkanoyl groups, such as formyl, acetyland the like; arylmethyl groups, such as benzyl (Bn), p-methoxybenzyl(PMB), 9-fluorenylmethyl (Fm), diphenylmethyl (benzhydryl, DPM) and thelike. Additionally, two hydroxyl groups can also be protected as analkylidene group, such as prop-2-ylidine, formed, for example, byreaction with a ketone, such as acetone.

General Synthetic Procedures

The compounds of this invention can be prepared from readily availablestarting materials using the following general methods and procedures orby using other information readily available to those of ordinary skillin the art. Although a particular embodiment of the present inventionmay be shown or described herein, those skilled in the art willrecognize that all embodiments or aspects of the present invention canbe prepared using the methods described herein or by using othermethods, reagents and starting materials known to those skilled in theart. It will also be appreciated that where typical or preferred processconditions (i.e., reaction temperatures, times, mole ratios ofreactants, solvents, pressures, etc.) are given, other processconditions can also be used unless otherwise stated. While the optimumreaction conditions may vary depending on the particular reactants orsolvent used, such conditions can be readily determined by one skilledin the art by routine optimization procedures.

Additionally, as will be apparent to those skilled in the art,conventional protecting groups may be necessary or desired to preventcertain functional groups from undergoing undesired reactions. Thechoice of a suitable protecting group for a particular functional groupas well as suitable conditions for protection and deprotection of suchfunctional groups are well-known in the art. Protecting groups otherthan those illustrated in the procedures described herein may be used,if desired. For example, numerous protecting groups, and theirintroduction and removal, are described in T. W. Greene and G. M. Wuts,Protecting Groups in Organic Synthesis, Third Edition, Wiley, New York,1999, and references cited therein.

By way of illustration, the compounds of this invention can be preparedby a process comprising:

(a) reacting a compound of formula 1:

or a salt thereof; with a compound of formula 2:

wherein X¹ represents a leaving group, and P¹ and P² each independentlyrepresent hydrogen or a hydroxyl-protecting group;

(b) reacting a compound of formula 3:

or salt thereof; wherein P³ represents hydrogen or an amino-protectinggroup, with a compound of formula 4:

wherein X² represents a leaving group, and P⁴ and P⁵ each independentlyrepresent hydrogen or a hydroxyl-protecting group;

(c) coupling a compound of formula 5:

with a compound of formula 6:

wherein X^(Qa) and X^(Qb) each independently represent functional groupsthat couple to form a group Q, P⁶ represents hydrogen or anamino-protecting group; and P⁷ and P⁸ each independently representhydrogen or a hydroxyl-protecting group;

(d) for a compound of formula I wherein R⁵ represents hydrogen, reactinga compound of formula 3 with a compound of formula 7:

or a hydrate thereof (e.g., a glyoxal), wherein P⁹ represents hydrogenor a hydroxyl-protecting group, in the presence of a reducing agent;

(e) reacting a compound of formula 1 with a compound of formula 8:

or a hydrate thereof, in the presence of a reducing agent, wherein P¹⁰and P¹¹ each independently represent hydrogen or a hydroxyl-protectinggroup; P¹² represents hydrogen or an amino-protecting group; and R^(4′)represents a residue that, together with the carbon to which it isattached, affords a group R⁴ upon completion of the reaction;

(f) reacting a compound of formula 9:

wherein X³ represents a leaving group, with a compound of formula 10:

wherein P¹³ and P¹⁴ each independently represent hydrogen or ahydroxyl-protecting group, and P¹⁵ represents hydrogen or anamino-protecting group;

(g) reacting a compound of formula 11:

with a reducing agent; wherein P¹⁶ represents hydrogen or anamino-protecting group; and P¹⁷ represents hydrogen or ahydroxyl-protecting group;

(h) for a compound of formula I in which E represents —C(O)NW^(a)W^(b),reacting a compound of formula 12:

wherein P¹⁸ and P¹⁹ each represents hydrogen or a hydroxyl-protectinggroup, with a compound of formula 13:

or

(i) reacting a compound of formula 14:

or a hydrate thereof; wherein R⁴″ represents a residue that, togetherwith the carbon to which it is attached, affords an R⁴ group uponcompletion of the reaction; with a compound of formula 10 in thepresence of a reducing agent;

and then removing any protecting group P¹, P², P¹, P⁴, P⁵, P⁶, P⁷, P⁸,P⁹, P¹¹, P¹², P¹³, P¹⁴, P¹⁵, P¹⁶, P¹⁷, P¹⁸ or P¹⁹ to provide a compoundof formula I; and optionally, forming a pharmaceutically acceptable saltthereof.

Generally, if a salt of one of the starting materials is used in theprocesses described above, such as an acid addition salt, the salt istypically neutralized before or during the reaction process. Thisneutralization reaction is typically accomplished by contacting the saltwith one molar equivalent of a base for each molar equivalent of acidaddition salt.

In process (a), i.e., the reaction between the compounds of formula 1and 2, the leaving group represented by X¹ can be, for example, halo,such as chloro, bromo or iodo, or a sulfonic ester group, such asmesylate or tosylate. The groups P¹ and P² can be, for example,trimethylsilyl and benzyl, respectively. This reaction is typicallyconducted in an inert diluent, such as acetonitrile, in the presence ofa base. For example, this reaction can be conducted in the presence of atertiary amine, such as diisopropylethylamine. Generally, this reactionis conducted at a temperature in the range of from 0° C. to 100° C.until the reaction is substantially complete. The reaction product isthen isolated using conventional procedures, such as extraction,recrystallization, chromatography and the like.

Compounds of formula 1 are generally known in the art or can be preparedfrom commercially available starting materials and reagents usingwell-known procedures. For example, compounds of formula 1 in whichW^(a) and W^(b) each represents hydrogen can be prepared using theprocedures described in U.S. Pat. No. 5,096,890, the disclosure of whichis incorporated herein by reference in its entirety.

By way of further illustration, compounds of formula 1 can be preparedby deprotecting a compound of formula 15:

wherein P²⁰ represents an amino-protecting group, such as a benzylgroup. Such benzyl groups are readily removed by reduction using, forexample, hydrogen or ammonium formate and a Group VIII metal catalyst,such as palladium. Optionally, this reaction is conducted in thepresence of an acid, such as formic acid, acetic acid, hydrochloricacid, hydrobromic acid, sulfuric acid and the like.

Compounds of formula 15 in which E is —C(O)NW^(a)W^(b) can be preparedby reacting a carboxylic acid of formula 16:

with an amine of formula 13 under amide bond forming conditions.Compounds of formula 16 may be prepared by hydrolyzing a compound offormula 15 in which E represents CN. Compounds of formula 15 can beprepared as described in U.S. Pat. No. 5,096,890.

Compounds of formula 2 can be prepared by various procedures describedherein or by procedures that are well-known to those skilled in the art.For example, the hydroxyl group of a compound of formula 25 below, canbe readily converted into a leaving group using well-known reagents andprocedures. By way of illustration, a hydroxyl group can be convertedinto a halo group using an inorganic acid halide, such as thionylchloride, phosphorous trichloride, phosphorous tribromide, phosphorousoxychloride and the like, or a halogen acid, such a hydrogen bromide.

In process (b), i.e., the reaction of a compound of formula 3 with acompound of formula 4, the leaving represented by X² can be, forexample, halo, such as chloro, bromo or iodo, or a sulfonic ester group,such as mesylate or tosylate. The groups P³, P⁴ and P⁵ can be, forexample, benzyl, tert-butyldimethylsilyl and benzyl, respectively. Thisreaction is typically conducted in the presence of a base, such assodium bicarbonate, and optionally in the presence of an alkali metaliodide, such as sodium iodide. Generally, this reaction is conducted inan inert diluent, such as tetrahydrofuran, at a temperature ranging from25° C. to 100° C. until the reaction is substantially complete. Thereaction product is then isolated using conventional procedures, such asextraction, recrystallization, chromatography and the like.

Compounds of formula 3 can be prepared by deprotecting a compound offormula 17:

wherein one or both of P²¹ and P²² independently represents a protectinggroup, such as tert-butoxycarbonyl or benzyl, and any remainderrepresents a hydrogen atom. For example, a tert-butoxycarbonyl group canbe removed by treating the protected compound with trifluoroacetic acid.

Compounds of formula 17 can be prepared by reacting a compound offormula 1 with a compound of formula 18:

wherein X³ represents a leaving group such as halo, such as chloro,bromo or iodo, or sulfonic ester group, such as mesylate or tosylate.This reaction is typically conducted by contacting a compound of formula1 with a compound of formula 18 in an inert diluent, such asacetonitrile, DMF or mixtures thereof, at a temperature ranging fromabout 0° C. to about 100° C. until the reaction is substantiallycomplete.

Alternatively, compounds of formula 3 can be obtained by reductiveamination of a compound of formula 14. The reductive amination can beperformed by reacting the compound of formula 14 with, for example,benzylamine in the presence of a reducing agent, including a borohydridereducing agent, such as sodium triacetoxyborohydride.

Compounds of formula 14 may be prepared by oxidizing the correspondingalcohol of formula 19:

using a suitable oxidizing agent, such as sulfur trioxide pyridinecomplex and dimethyl sulfoxide. This oxidation reaction is typicallyconducted in an inert diluent, such as dichloromethane, the presence ofa tertiary amine, such as diisopropylethylamine, at a temperatureranging from about −20° C. to about 25° C.

Compounds of formula 19 can be prepared by reacting a compound offormula 1 with a compound of formula 20:

wherein X⁴ represents a leaving group such as halo, such as chloro,bromo or iodo, or a sulfonic ester group, such as mesylate or tosylate.

Compounds of formula 4 can be prepared by reacting a compound of formula21:

with a reducing agent, such as borane. If desired, such a reduction canbe performed in the presence of a chiral catalyst to provide compoundsof formula 4 in chiral form. For example, compounds of formula 21 can bereduced in the presence of a chiral catalyst formed from(R)-(+)-α,α-diphenyl-2-pyrrolidinemethanol and trimethylboroxine; oralternatively, from (S)-(−)-α,α-diphenyl-2-pyrrolidinemethanol andtrimethylboroxine. The resulting hydroxyl group can then be protectedwith a hydroxyl-protecting group, P⁴, by reaction with, for example,tert-butyldimethylsilyl trifluoromethanesulfonate.

Compounds of formula 21 in which X² represents a bromine atom can beprepared by reacting a compound of formula 22:

with bromine in the presence of a Lewis acid, such as boron trifluoridediethyl etherate. Compounds of formula 22 are well-known in the art orcan be prepared by well-known procedures using commercially availablestarting materials and reagents.

Referring to process (c), i.e., the reaction of a compound of formula 5with a compound of formula 6, it will be appreciated that the groupsX^(Qa) and X^(Qb) should be selected so as to afford the desired group Qupon completion of the reaction. For example, when the desired group Qis an amide group, i.e., —N(Q^(a))C(O)— or —C(O)N(Q^(b)), one of X^(Qa)and X^(Qb) can be an amine group (i.e., —NHQ^(a) or —NHQ^(b)) and theother can be a carboxyl group (i.e., —COOH) or a reactive derivativethereof (such as acyl halide, such as an acyl chloride or acyl bromide).The groups P⁶, P⁷ and P⁸ can be, for example, benzyl, trimethylsilyl andbenzyl, respectively. When Q is an amide group, the reaction can beperformed under conventional amide coupling conditions. Similarly, whenthe desired group Q is a sulfonamide, i.e., —N(Q^(c))S(O)₂— or—S(O)₂N(Q^(d))-, one of X^(Qa) and X^(Qb) can be an amine group,—NHQ^(c) or —NHQ^(d) and the other can be a sulfonyl halide group (suchas sulfonyl chloride or sulfonyl bromide).

Compounds of formula 5 can be prepared by reacting a compound of formula1 with a compound of formula 23:

wherein X⁵ represents a leaving group including halo, such as chloro,bromo or iodo, and a sulfonic ester group, such as mesylate or tosylate;and X^(Qa′) represents X^(Qa), such as a carboxyl group or an aminogroup NHQ^(a), or a protected derivative thereof, such as a(1-6C)alkoxycarbonylamino group or a tert-butoxycarbonylamino group.This reaction is typically conducted by a method analogous to that usedto prepare compounds of formula 3, followed by removing any protectinggroup in X^(Qa′).

Compounds of formula 6 can be prepared by reacting a compound of formula4 with a compound of formula 24:

wherein X⁶ represents a leaving group including halo, such as chloro,bromo or iodo, and a sulfonic ester group, such as mesylate or tosylate;and X^(Qb′) represents X^(Qb), such as a carboxyl group or an aminogroup NHQ^(b), or a protected derivative thereof, such as a(1-6C)alkoxycarbonyl group or a tert-butoxycarbonylamino group. Thisreaction is typically conducted by a method analogous to that used toprepare compounds of formula 3, followed by removing any protectinggroup in X^(Qb′).

Referring to process (d), i.e., the reaction of a compound of formula 3with a compound of formula 7, any suitable reducing agent may be used inthis reaction. For example, the reducing agent can be hydrogen in thepresence of a Group VIII metal catalyst, such as palladium on carbon; ora metal hydride reagent, such as sodium triacetoxyborohydride. The groupP⁷ can be, for example, benzyl. This reaction is typically conducted inan inert diluent and a protic solvent, such as a mixture ofdichloroethane and methanol, at a temperature in the range of from 0° C.to 100° C. until the reaction is substantially complete.

Compounds of formula 7 in the form of a hydrate can be prepared byconventional procedures, for example, by dibrominating a compound offormula 21 (where X² in this case can also be hydrogen), and thenhydrolyzing the resulting dibromide to form a glyoxal or a hydratethereof. For example, a compound of formula 21 can be reacted withhydrogen bromide and then hydrolyzed with water to form thecorresponding glyoxal hydrate.

Referring to process (e), i.e., the reaction of a compound of formula 1with a compound of formula 8, any suitable reducing agent may be used inthis reaction. For example, the reducing agent may be hydrogen in thepresence of a Group VIII metal catalyst, such as palladium on carbon; ora metal hydride reagent, such as sodium triacetoxyborohydride. Thegroups P¹⁰, P¹¹ and P¹² can be, for example, trimethylsilyl, benzyl andbenzyl, respectively. Typically, this reduction reaction is conducted inan inert diluent and a protic solvent, such as dichloroethane andmethanol, at a temperature in the range of from 0° C. to 100° C. untilthe reaction is substantially complete.

Compounds of formula 8 may be prepared by oxidizing a compound offormula 25:

using any suitable oxidizing agent, such as sulfur trioxide pyridinecomplex and dimethyl sulfoxide. This reaction is typically conducted inthe presence of a tertiary amine, such as diisopropylethylamine, at atemperature in the range of from about −20° C. to about 25° C. until theoxidation is substantially complete.

Compounds of formula 25 can be prepared by reacting a compound offormula 10 with a compound of formula 26:

wherein X⁷ represents a leaving group including halo, such as chloro,bromo or iodo, and a sulfonic ester group, such as mesylate or tosylate.

Referring to process (f), i.e., the reaction of a compound of formula 9with a compound of formula 10, the leaving group represented by X³ canbe, for example, halo, such as chloro, bromo or iodo, or a sulfonicester group, such as mesylate or tosylate. The groups P¹³, P¹⁴ and P¹⁵can be, for example, trimethylsilyl, benzyl and benzyl, respectively.This reaction is typically conducted an inert diluent, such asacetonitrile, in the presence of a suitable base. For example, thisreaction can be conducted in the presence of a tertiary amine, such asdiisopropylethylamine. Generally, this reaction is conducted at atemperature in the range of from 0° C. to 100° C. until the reaction issubstantially complete.

Compounds of formula 9 can be prepared by steps analogous to those ofmethods (a) to (e) herein, starting from a compound of formula 1.Additionally, compounds of formula 10 can be prepared from compounds offormula 4 by reaction with an amine of formula P¹⁵NH₂.

Referring to process (g), representative examples for P¹⁶ and P¹⁷ arebenzyl or tert-butyldimethylsilyl. This reduction reaction can beconducted using any suitable reducing agent including borohydrides, suchas sodium borohydride. Any suitable solvent or diluent may be employed,such as N,N-dimethylformamide. This reaction is typically conducted at atemperature in the range of from 0° C. to 100° C. until the reaction issubstantially complete.

Compounds of formula 11 can be prepared by reacting a compound offormula 3 with a compound of formula 21. This reaction is convenientlyperformed in the presence of a base, such as potassium carbonate.

Referring to process (h), the hydroxyl-protecting groups represented byP¹⁸ and P¹⁹ can be, for example, trimethylsilyl and benzyl. Thisreaction is typically performed under conventional amide bond formingconditions.

Compounds of formula 12 can be prepared using the methods as describedherein by employing a starting material in which E is —C(O)OW^(c).Alternatively, such compounds can be prepared by hydrolyzing a compoundcorresponding to 12 in which the —COOH group is —CN.

Referring to process (i), i.e., the reaction of a compound of formula 14with a compound of formula 10, any suitable reducing agent may be usedin this reaction. For example, the reducing agent may be hydrogen in thepresence of a Group VIII metal catalyst, such as palladium on carbon; ora metal hydride reagent, such as sodium triacetoxyborohydride. Thegroups P¹³, P¹⁴ and P¹⁵ can be, for example, tert-butyldimethylsilyl,benzyl and benzyl, respectively. Typically, this reduction reaction isconducted in an inert diluent and a protic solvent, such asdichloroethane and methanol, at a temperature in the range of from 0° C.to 100° C. until the reaction is substantially complete.

Compounds of formula 14 are readily prepared by oxidation of thecorresponding alcohol or by hydrolysis of the corresponding acetal. Anysuitable oxidizing agent may be employed in this reaction to provide thealdehyde, such as sulfur trioxide pyridine complex and dimethylsulfoxide. The acetal may be hydrolyzed under conventional conditionsusing aqueous acid to provide the aldehyde.

Additionally, compounds of formula I in which R⁶ and R⁷ together form—NR^(7g)C(O)—CR^(7h)R^(7i)—CR^(7j)R^(7k)— or—CR^(7l)R^(7m)—CR^(7n)R^(7o)—C(O)—NR^(7p)— may be prepared by reducing acorresponding compound of formula I in which R⁶ and R⁷ together form—NR^(7a)C(O)—CR^(7b)═CR^(7c)— or —CR^(7d)═CR^(7e)—C(O)—NR^(7f)—, forexample by catalytic hydrogenation.

Further details regarding specific reaction conditions and otherprocedures for preparing representative compounds of this invention orintermediates thereof are described in the Examples set forth below.

Pharmaceutical Compositions and Formulations

The compounds of this invention are typically administered to a patientin the form of a pharmaceutical composition or formulation. Suchpharmaceutical compositions may be administered to the patient by anyacceptable route of administration including, but not limited to,inhaled, oral, nasal, topical (including transdermal) and parenteralmodes of administration. It will be understood that any form of thecompounds of this invention, (i.e., free base, pharmaceuticallyacceptable salt, solvate, etc.) that is suitable for the particular modeof administration can be used in the pharmaceutical compositionsdiscussed herein.

Accordingly, in one of its compositions aspects, this invention isdirected to a pharmaceutical composition comprising a pharmaceuticallyacceptable carrier or excipient and a therapeutically effective amountof a compound of formula I, or a pharmaceutically acceptable saltthereof. Optionally, such pharmaceutical compositions may contain othertherapeutic and/or formulating agents if desired.

The pharmaceutical compositions of this invention typically contain atherapeutically effective amount of a compound of the present inventionor a pharmaceutically acceptable salt thereof. Typically, suchpharmaceutical compositions will contain from about 0.01 to about 95% byweight of the active agent; including, from about 0.01 to about 30% byweight; such as from about 0.01 to about 10% by weight of the activeagent.

Any conventional carrier or excipient may be used in the pharmaceuticalcompositions of this invention. The choice of a particular carrier orexcipient, or combinations of carriers or excipients, will depend on themode of administration being used to treat a particular patient or typeof medical condition or disease state. In this regard, the preparationof a suitable pharmaceutical composition for a particular mode ofadministration is well within the scope of those skilled in thepharmaceutical arts. Additionally, the ingredients for such compositionsare commercially available from, for example, Sigma, P.O. Box 14508, St.Louis, Mo. 63178. By way of further illustration, conventionalformulation techniques are described in Remington: The Science andPractice of Pharmacy, 20^(th) Edition, Lippincott Williams & White,Baltimore, Md. (2000); and H. C. Ansel et al., Pharmaceutical DosageForms and Drug Delivery Systems, 7^(th) Edition, Lippincott Williams &White, Baltimore, Md. (1999).

Representative examples of materials which can serve as pharmaceuticallyacceptable carriers include, but are not limited to, the following: (1)sugars, such as lactose, glucose and sucrose; (2) starches, such as cornstarch and potato starch; (3) cellulose, and its derivatives, such assodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate;(4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8)excipients, such as cocoa butter and suppository waxes; (9) oils, suchas peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil,corn oil and soybean oil; (10) glycols, such as propylene glycol; (11)polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol;(12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14)buffering agents, such as magnesium hydroxide and aluminum hydroxide;(15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18)Ringer's solution; (19) ethyl alcohol; (20) phosphate buffer solutions;(21) compressed propellant gases, such as chlorofluorocarbons andhydrofluorocarbons; and (22) other non-toxic compatible substancesemployed in pharmaceutical compositions.

The pharmaceutical compositions of this invention are typically preparedby thoroughly and intimately mixing or blending a compound of theinvention with a pharmaceutically acceptable carrier and one or moreoptional ingredients. If necessary or desired, the resulting uniformlyblended mixture can then be shaped or loaded into tablets, capsules,pills, canisters, cartridges, dispensers and the like using conventionalprocedures and equipment.

In one embodiment, the pharmaceutical compositions of this invention aresuitable for inhaled administration. Suitable pharmaceuticalcompositions for inhaled administration will typically be in the form ofan aerosol or a powder. Such compositions are generally administeredusing well-known delivery devices, such as a nebulizer inhaler, ametered-dose inhaler (MDI), a dry powder inhaler (DPI) or a similardelivery device.

In a specific embodiment of this invention, the pharmaceuticalcomposition comprising the active agent is administered by inhalationusing a nebulizer inhaler. Such nebulizer devices typically produce astream of high velocity air that causes the pharmaceutical compositioncomprising the active agent to spray as a mist that is carried into thepatient's respiratory tract. Accordingly, when formulated for use in anebulizer inhaler, the active agent is typically dissolved in a suitablecarrier to form a solution. Alternatively, the active agent can bemicronized and combined with a suitable carrier to form a suspension ofmicronized particles of respirable size, where micronized is typicallydefined as having about 90% or more of the particles with a diameter ofless than about 10 μm. Suitable nebulizer devices are providedcommercially, for example, by PARI GmbH (Starnberg, German). Othernebulizer devices include Respimat (Boehringer Ingelheim) and thosedisclosed, for example, in U.S. Pat. No. 6,123,068 and WO 97/12687.

A representative pharmaceutical composition for use in a nebulizerinhaler comprises an isotonic aqueous solution comprising from about0.05 μg/mL to about 10 mg/mL of a compound of formula I or apharmaceutically acceptable salt or solvate or stereoisomer thereof.

In another specific embodiment of this invention, the pharmaceuticalcomposition comprising the active agent is administered by inhalationusing a dry powder inhaler. Such dry powder inhalers typicallyadminister the active agent as a free-flowing powder that is dispersedin a patient's air-stream during inspiration. In order to achieve a freeflowing powder, the active agent is typically formulated with a suitableexcipient such as lactose or starch.

A representative pharmaceutical composition for use in a dry powderinhaler comprises dry lactose having a particle size between about 1 μmand about 100 μm and micronized particles of a compound of formula I, ora pharmaceutically acceptable salt or solvate or stereoisomer thereof.

Such a dry powder formulation can be made, for example, by combining thelactose with the active agent and then dry blending the components.Alternatively, if desired, the active agent can be formulated without anexcipient. The pharmaceutical composition is then typically loaded intoa dry powder dispenser, or into inhalation cartridges or capsules foruse with a dry powder delivery device.

Examples of dry powder inhaler delivery devices include Diskhaler(GlaxoSmithKline, Research Triangle Park, N.C.) (see, e.g., U.S. Pat.No. 5,035,237); Diskus (GlaxoSmithKline) (see, e.g., U.S. Pat. No.6,378,519; Turbuhaler (AstraZeneca, Wilmington, Del.) (see, e.g., U.S.Pat. No. 4,524,769); Rotahaler (GlaxoSmithKline) (see, e.g., U.S. Pat.No. 4,353,365) and Handihaler (Boehringer Ingelheim). Further examplesof suitable DPI devices are described in U.S. Pat. Nos. 5,415,162,5,239,993, and 5,715,810 and references cited therein.

In yet another specific embodiment of this invention, the pharmaceuticalcomposition comprising the active agent is administered by inhalationusing a metered-dose inhaler. Such metered-dose inhalers typicallydischarge a measured amount of the active agent or a pharmaceuticallyacceptable salt thereof using compressed propellant gas. Accordingly,pharmaceutical compositions administered using a metered-dose inhalertypically comprise a solution or suspension of the active agent in aliquefied propellant. Any suitable liquefied propellant may be employedincluding chlorofluorocarbons, such as CCl₃F, and hydrofluoroalkanes(HFAs), such as 1,1,1,2-tetrafluoroethane (HFA 134a) and1,1,1,2,3,3,3-heptafluoro-n-propane, (HFA 227). Due to concerns aboutchlorofluorocarbons affecting the ozone layer, formulations containingHFAs are generally preferred. Additional optional components of HFAformulations include co-solvents, such as ethanol or pentane, andsurfactants, such as sorbitan trioleate, oleic acid, lecithin, andglycerin. See, for example, U.S. Pat. No. 5,225,183, EP 0717987 A2, andWO 92/22286.

A representative pharmaceutical composition for use in a metered-doseinhaler comprises from about 0.01% to about 5% by weight of a compoundof formula 1, or a pharmaceutically acceptable salt or solvate orstereoisomer thereof; from about 0% to about 20% by weight ethanol; andfrom about 0% to about 5% by weight surfactant; with the remainder beingan HFA propellant.

Such compositions are typically prepared by adding chilled orpressurized hydrofluoroalkane to a suitable container containing theactive agent, ethanol (if present) and the surfactant (if present). Toprepare a suspension, the active agent is micronized and then combinedwith the propellant. The formulation is then loaded into an aerosolcanister, which forms a portion of a metered-dose inhaler device.Examples of metered-dose inhaler devices developed specifically for usewith HFA propellants are provided in U.S. Pat. Nos. 6,006,745 and6,143,277. Alternatively, a suspension formulation can be prepared byspray drying a coating of surfactant on micronized particles of theactive agent. See, for example, WO 99/53901 and WO 00/61108.

For additional examples of processes of preparing respirable particles,and formulations and devices suitable for inhalation dosing see U.S.Pat. Nos. 6,268,533, 5,983,956, 5,874,063, and 6,221,398, and WO99/55319 and WO 00/30614.

In another embodiment, the pharmaceutical compositions of this inventionare suitable for oral administration. Suitable pharmaceuticalcompositions for oral administration may be in the form of capsules,tablets, pills, lozenges, cachets, dragees, powders, granules; or as asolution or a suspension in an aqueous or non-aqueous liquid; or as anoil-in-water or water-in-oil liquid emulsion; or as an elixir or syrup;and the like; each containing a predetermined amount of a compound ofthe present invention as an active ingredient.

When intended for oral administration in a solid dosage form (i.e., ascapsules, tablets, pills and the like), the pharmaceutical compositionsof this invention will typically comprise a compound of the presentinvention as the active ingredient and one or more pharmaceuticallyacceptable carriers, such as sodium citrate or dicalcium phosphate.Optionally or alternatively, such solid dosage forms may also comprise:(1) fillers or extenders, such as starches, lactose, sucrose, glucose,mannitol, and/or silicic acid; (2) binders, such ascarboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone,sucrose and/or acacia; (3) humectants, such as glycerol; (4)disintegrating agents, such as agar-agar, calcium carbonate, potato ortapioca starch, alginic acid, certain silicates, and/or sodiumcarbonate; (5) solution retarding agents, such as paraffin; (6)absorption accelerators, such as quaternary ammonium compounds; (7)wetting agents, such as cetyl alcohol and/or glycerol monostearate; (8)absorbents, such as kaolin and/or bentonite clay; (9) lubricants, suchas talc, calcium stearate, magnesium stearate, solid polyethyleneglycols, sodium lauryl sulfate, and/or mixtures thereof; (10) coloringagents; and (11) buffering agents.

Release agents, wetting agents, coating agents, sweetening, flavoringand perfuming agents, preservatives and antioxidants can also be presentin the pharmaceutical compositions of this invention. Examples ofpharmaceutically acceptable antioxidants include: (1) water-solubleantioxidants, such as ascorbic acid, cysteine hydrochloride, sodiumbisulfate, sodium metabisulfate sodium sulfite and the like; (2)oil-soluble antioxidants, such as ascorbyl palmitate, butylatedhydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propylgallate, alpha-tocopherol, and the like; and (3) metal-chelating agents,such as citric acid, ethylencdiamine tetraacetic acid (EDTA), sorbitol,tartaric acid, phosphoric acid, and the like. Coating agents fortablets, capsules, pills and like, include those used for entericcoatings, such as cellulose acetate phthalate (CAP), polyvinyl acetatephthalate (PVAP), hydroxypropyl methylcellulose phthalate, methacrylicacid-methacrylic acid ester copolymers, cellulose acetate trimellitate(CAT), carboxymethyl ethyl cellulose (CMEC), hydroxypropyl methylcellulose acetate succinate (HPMCAS), and the like.

If desired, the pharmaceutical compositions of the present invention mayalso be formulated to provide slow or controlled release of the activeingredient using, by way of example, hydroxypropyl methyl cellulose invarying proportions; or other polymer matrices, liposomes and/ormicrospheres.

In addition, the pharmaceutical compositions of the present inventionmay optionally contain opacifying agents and may be formulated so thatthey release the active ingredient only, or preferentially, in a certainportion of the gastrointestinal tract, optionally, in a delayed manner.Examples of embedding compositions which can be used include polymericsubstances and waxes. The active ingredient can also be inmicro-encapsulated form, if appropriate, with one or more of theabove-described excipients.

Suitable liquid dosage forms for oral administration include, by way ofillustration, pharmaceutically acceptable emulsions, microemulsions,solutions, suspensions, syrups and elixirs. Such liquid dosage formstypically comprise the active ingredient and an inert diluent, such as,for example, water or other solvents, solubilizing agents andemulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate,ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol,1,3-butylene glycol, oils (esp., cottonseed, groundnut, corn, germ,olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol,polyethylene glycols and fatty acid esters of sorbitan, and mixturesthereof. Suspensions, in addition to the active ingredient, may containsuspending agents such as, for example, ethoxylated isostearyl alcohols,polyoxyethylene sorbitol and sorbitan esters, microcrystallinecellulose, aluminium metahydroxide, bentonite, agar-agar and tragacanth,and mixtures thereof.

When intended for oral administration, the pharmaceutical compositionsof this invention are preferably packaged in a unit dosage form. Theterm “unit dosage form” means a physically discrete unit suitable fordosing a patient, i.e., each unit containing a predetermined quantity ofactive agent calculated to produce the desired therapeutic effect eitheralone or in combination with one or more additional units. For example,such unit dosage forms may be capsules, tablets, pills, and the like.

The compounds of this invention can also be administered transdermallyusing known transdermal delivery systems and excipients. For example, acompound of this invention can be admixed with permeation enhancers,such as propylene glycol, polyethylene glycolm monolaurate,azacycloalkan-2-ones and the like, and incorporated into a patch orsimilar delivery system. Additional excipients including gelling agents,emulsifiers and buffers, may be used in such transdermal compositions ifdesired.

The pharmaceutical compositions of this invention may also contain othertherapeutic agents that are co-administered with a compound of formulaI, or pharmaceutically acceptable salt or solvate or stereoisomerthereof. For example, the pharmaceutical compositions of this inventionmay further comprise one or more therapeutic agents selected from otherbronchodilators (e.g., PDE₃ inhibitors, adenosine 2b modulators and β₂adrenergic receptor agonists); anti-inflammatory agents (e.g. steroidalanti-inflammatory agents, such as corticosteroids; non-steroidalanti-inflammatory agents (NSAIDs), and PDE₄ inhibitors); othermuscarinic receptor antagonists (i.e., antichlolinergic agents);antiinfective agents (e.g. Gram positive and Gram negative antibioticsor antivirals); antihistamines; protease inhibitors; and afferentblockers (e.g., D₂ agonists and neurokinin modulators). The othertherapeutic agents can be used in the form of pharmaceuticallyacceptable salts or solvates. Additionally, if appropriate, the othertherapeutic agents can be used as optically pure stereoisomers.

Representative β₂ adrenergic receptor agonists that can be used incombination with, and in addition to, the compounds of this inventioninclude, but are not limited to, salmeterol, salbutamol, formoterol,salmefamol, fenoterol, terbutaline, albuterol, isoetharine,metaproterenol, bitolterol, pirbuterol, levalbuterol and the like, orpharmaceutically acceptable salts thereof. Other β₂ adrenergic receptoragonists that can be used in combination with the compounds of thisinvention include, but are not limited to,3-(4-{[6-({(2R)-2-hydroxy-2-[4-hydroxy-3-(hydroxymethyl)-phenyl]ethyl}amino)-hexyl]oxy}butyl)benzenesulfonamideand3-(-3-{[7-({(2R)-2-hydroxy-2-[4-hydroxy-3-(hydroxymethyl)phenyl]ethyl}-amino)heptyl]oxy}-propyl)benzenesulfonamideand related compounds disclosed in WO 02/066422, published on Aug. 29,2002;3-[3-(4-{[6-([(2R)-2-hydroxy-2-[4-hydroxy-3-(hydroxymethyl)phenyl]ethyl}amino)hexyl]oxy}butyl)-phenyl]imidazolidine-2,4-dioneand related compounds disclosed in WO 02/070490, published Sep. 12,2002;3-(4-{[6-({(2R)-2-[3-(formylamino)-4-hydroxyphenyl]-2-hydroxyethyl}amino)hexyl]oxy}butyl)-benzenesulfonamide,3-(4-{[6-({(2S)-2-[3-(formylamino)-4-hydroxyphenyl]-2-hydroxyethyl}amino)hexyl]oxy}butyl)-benzenesulfonamide,3-(4-{[6-({(2R/S)-2-[3-(formylamino)-4-hydroxyphenyl]-2-hydroxyethyl}amino)hexyl]oxy}butyl)-benzenesulfonamide,N-(tert-butyl)-3-(4-{[6-({(2R)-2-[3-(formylamino)-4-hydroxyphenyl]-2-hydroxyethyl}amino)hexyl]-oxy}butyl)benzenesulfonamide,N-(tert-butyl)-3-(4-{[6-({(2S)-2-[3-(formylamino)-4-hydroxyphenyl]-2-hydroxyethyl}amino)-hexyl]oxy}butyl)-benzenesulfonamide,N-(tert-butyl)-3-(4-{[6-({(2R/S)-2-[3-(formylamino)-4-hydroxyphenyl]-2-hydroxyethyl}amino)hexyl]-oxy}butyl)benzenesulfonamideand related compounds disclosed in WO 02/076933, published on Oct. 3,2002;4-{(1R)-2-[(6-{2-[(2,6-dichlorobenzyl)oxy]ethoxy}hexyl)amino]-1-hydroxyethyl}-2-(hydroxymethyl)phenoland related compounds disclosed in WO 03/024439, published on Mar. 27,2003; and pharmaceutically acceptable salts thereof. When employed, theβ₂-adrenoreceptor agonist will be present in the pharmaceuticalcomposition in a therapeutically effective amount. Typically, theβ₂-adrenoreceptor agonist will be present in an amount sufficient toprovide from about 0.05 μg to about 500 μg per dose.

Representative steroidal anti-inflammatory agents that can be used incombination with the compounds of this invention include, but are notlimited to, methyl prednisolone, prednisolone, dexamethasone,fluticasone propionate,6,9-difluoro-17-[(2-furanylcarbonyl)oxy]-11-hydroxy-16-methyl-3-oxoandrosta-1,4-diene-17-carbothioicacid S-fluoromethyl ester,6,9-difluoro-11-hydroxy-16-methyl-3-oxo-17-propionyloxy-androsta-1,4-diene-17-carbothioicacid S-(2-oxo-tetrahydrofuran-3S-yl) ester, beclomethasone esters (e.g.the 17-propionate ester or the 17,21-dipropionate ester), budesonide,flunisolide, mometasone esters (e.g. the furoate ester), triamcinoloneacetonide, rofleponide, ciclesonide, butixocort propionate, RPR-106541,ST-126 and the like, or pharmaceutically-acceptable salts thereof. Whenemployed, the steroidal anti-inflammatory agent will be present in thepharmaceutical composition in a therapeutically effective amount.Typically, the steroidal anti-inflammatory agent will be present in anamount sufficient to provide from about 0.05 μg to about 500 μg perdose.

Other suitable combinations include, for example, otheranti-inflammatory agents, e.g., NSAIDs (such as sodium cromoglycate;nedocromil sodium; phosphodiesterase (PDE) inhibitors (e.g.theophylline, PDE4 inhibitors or mixed PDE3/PDE4 inhibitors);leukotriene antagonists (e.g. monteleukast); inhibitors of leukotrienesynthesis; iNOS inhibitors; protease inhibitors, such as tryptase andelastase inhibitors; beta-2 integrin antagonists and adenosine receptoragonists or antagonists (e.g. adenosine 2a agonists); cytokineantagonists (e.g. chemokine antagonists such as, an interleukin antibody(IL antibody), specifically, an IL-4 therapy, an IL-13 therapy, or acombination thereof); or inhibitors of cytokine synthesis.

For example, representative phosphodiesterase-4 (PDE4) inhibitors ormixed PDE3/PDE4 inhibitors that can be used in combination with thecompounds of this invention include, but are not limited to cis4-cyano-4-(3-cyclopentyloxy-4-methoxyphenyl)cyclohexan-1-carboxylicacid,2-carbomethoxy-4-cyano-4-(3-cyclopropylmethoxy-4-difluoromethoxyphenyl)cyclohexan-1-one;cis-[4-cyano-4-(3-cyclopropylmethoxy-4-difluoromethoxyphenyl)cyclohexan-1-ol];cis-4-cyano-4-[3-(cyclopentyloxy)-4-methoxyphenyl]cyclohexane-1-carboxylicacid and the like, or pharmaceutically acceptable salts thereof. Otherrepresentative PDE4 or mixed PDE4/PDE3 inhibitors include AWD-12-281(elbion); NCS-613 (INSERM); D-4418 (Chiroscience and Schering-Plough);CI-1018 or PD-168787 (Pfizer); benzodioxole compounds disclosed inWO99/16766 (Kyowa Hakko); K-34 (Kyowa Hakko); V-11294A (Napp);roflumilast (Byk-Gulden); pthalazinone compounds disclosed in WO99/47505(Byk-Gulden); Pumafentrine (Byk-Gulden, now Altana); arofylline(Almirall-Prodesfarma); VM554/UM565 (Vernalis); T-440 (Tanabe Seiyaku);and T2585 (Tanabe Seiyaku).

Representative muscarinic antagonists (i.e., anticholinergic agents)that can be used in combination with, and in addition to, the compoundsof this invention include, but are not limited to, atropine, atropinesulfate, atropine oxide, methylatropine nitrate, homatropinehydrobromide, hyoscyamine (d, l) hydrobromide, scopolamine hydrobromide,ipratropium bromide, oxitropium bromide, tiotropium bromide,methantheline, propantheline bromide, anisotropine methyl bromide,clidinium bromide, copyrrolate (Robinul), isopropamide iodide,mepenzolate bromide, tridihexethyl chloride (Pathilone), hexocycliummethylsulfate, cyclopentolate hydrochloride, tropicamide,trihexyphenidyl hydrochloride, pirenzepine, telenzepine, AF-DX 116 andmethoctramine and the like, or a pharmaceutically acceptable saltthereof; or, for those compounds listed as a salt, alternatepharmaceutically acceptable salt thereof.

Representative antihistamines (i.e., H₁-receptor antagonists) that canbe used in combination with the compounds of this invention include, butare not limited to, ethanolamines, such as carbinoxamine maleate,clemastine fumarate, diphenylhydramine hydrochloride and dimenhydrinate;ethylenediamines, such as pyrilamine amleate, tripelennaminehydrochloride and tripelennamine citrate; alkylamines, such aschlorpheniramine and acrivastine; piperazines, such as hydroxyzinehydrochloride, hydroxyzine pamoate, cyclizine hydrochloride, cyclizinelactate, meclizine hydrochloride and cetirizine hydrochloride;piperidines, such as astemizole, levocabastine hydrochloride, loratadineor its descarboethoxy analogue, terfenadine and fexofenadinehydrochloride; azelastine hydrochloride; and the like, or apharmaceutically acceptable salt thereof; or, for those compounds listedas a salt, alternate pharmaceutically acceptable salt thereof.

Suitable doses for the other therapeutic agents administered incombination with a compound of the invention are in the range of about0.05 μg/day to about 100 mg/day.

The following formulations illustrate representative pharmaceuticalcompositions of the present invention:

FORMULATION EXAMPLE A

A dry powder for administration by inhalation is prepared as follows:

Ingredients Amount Compound of the invention 0.2 mg Lactose  25 mg

-   -   Representative Procedure: The compound of the invention is        micronized and then blended with lactose. This blended mixture        is then loaded into a gelatin inhalation cartridge. The contents        of the cartridge are administered using a powder inhaler.

FORMULATION EXAMPLE B

A dry powder formulation for use in a dry powder inhalation device isprepared as follows:

Representative Procedure: A pharmaceutical composition is preparedhaving a bulk formulation ratio of micronized compound of the inventionto lactose of 1:200. The composition is packed into a dry powderinhalation device capable of delivering between about 10 μg and about100 μg of the compound of the invention per dose.

FORMULATION EXAMPLE C

A dry powder for administration by inhalation in a metered dose inhaleris prepared as follows:

Representative Procedure: A suspension containing 5 wt. % of a compoundof the invention and 0.1 wt. % lecithin is prepared by dispersing 10 gof the compound of the invention as micronized particles with mean sizeless than 10 μm in a solution formed from 0.2 g of lecithin dissolved in200 mL of demineralized water. The suspension is spray dried and theresulting material is micronized to particles having a mean diameterless than 1.5 μm. The particles are loaded into cartridges withpressurized 1,1,1,2-tetrafluoroethane.

FORMULATION EXAMPLE D

A pharmaceutical composition for use in a metered dose inhaler isprepared as follows:

Representative Procedure: A suspension containing 5% compound of theinvention, 0.5% lecithin, and 0.5% trehalose is prepared by dispersing 5g of active ingredient as micronized particles with mean size less than10 m in a colloidal solution formed from 0.5 g of trehalose and 0.5 g oflecithin dissolved in 100 mL of demineralized water. The suspension isspray dried and the resulting material is micronized to particles havinga mean diameter less than 1.5 μm. The particles are loaded intocanisters with pressurized 1,1,1,2-tetrafluoroethane.

FORMULATION EXAMPLE E

A pharmaceutical composition for use in a nebulizer inhaler is preparedas follows:

Representative Procedure: An aqueous aerosol formulation for use in anebulizer is prepared by dissolving 0.1 mg of the compound of theinvention in 1 mL of a 0.9% sodium chloride solution acidified withcitric acid. The mixture is stirred and sonicated until the activeingredient is dissolved. The pH of the solution is adjusted to a valuein the range of from 3 to 8 by the slow addition of NaOH.

FORMULATION EXAMPLE F

Hard gelatin capsules for oral administration are prepared as follows:

Ingredients Amount Compound of the invention 250 mg Lactose(spray-dried) 200 mg Magnesium stearate  10 mg

Representative Procedure: The ingredients are thoroughly blended andthen loaded into a hard gelatin capsule (460 mg of composition percapsule).

FORMULATION EXAMPLE G

A suspension for oral administration is prepared as follows:

Ingredients Amount Compound of the invention 1.0 g Fumaric acid 0.5 gSodium chloride 2.0 g Methyl paraben 0.15 g Propyl paraben 0.05 gGranulated sugar 25.5 g Sorbitol (70% solution) 12.85 g Veegum k(Vanderbilt Co.) 1.0 g Flavoring 0.035 mL Colorings 0.5 mg Distilledwater q.s. to 100 mL

-   -   Representative Procedure: The ingredients are mixed to form a        suspension containing 100 mg of active ingredient per 10 mL of        suspension.

FORMULATION EXAMPLE H

An injectable formulation is prepared as follows:

Ingredients Amount Compound of the invention 0.2 g Sodium acetate buffersolution (0.4 M) 2.0 mL HCl (0.5 N) or NaOH (0.5 N) q.s. to pH 4 Water(distilled, sterile) q.s. to 20 mL

-   -   Representative Procedure: The above ingredients are blended and        the pH is adjusted to 4±0.5 using 0.5 N HCl or 0.5 N NaOH.

Utility

The compounds of this invention possess both β₂ adrenergic receptoragonist and muscarinic receptor antagonist activity and therefore, suchcompounds are useful for treating medical conditions mediated by β₂adrenergic receptors or muscarinic receptors, i.e., medical conditionsthat are ameliorated by treatment with a β₂ adrenergic receptor agonistor a muscarinic receptor antagonist. Such medical conditions include, byway of example, pulmonary disorders or diseases associated withreversible airway obstruction, such as chronic obstructive pulmonarydisease (e.g., chronic and wheezy bronchitis and emphysema), asthma,pulmonary fibrosis and the like. Other conditions which may be treatedinclude premature labor, depression, congestive heart failure, skindiseases (e.g., inflammatory, allergic, psoriatic and proliferative skindiseases, conditions where lowering peptic acidity is desirable (e.g.,peptic and gastric ulceration) and muscle wasting disease.

Accordingly, in one embodiment, this invention is directed to a methodfor treating a pulmonary disorder, the method comprising administeringto a patient in need of treatment a therapeutically effective amount ofa compound of formula I or a pharmaceutically acceptable salt or solvateor stereoisomer thereof. When used to treat a pulmonary disorder, thecompounds of this invention will typically be administered by inhalationin multiple doses per day, in a single daily dose or a single weeklydose. Generally, the dose for treating a pulmonary disorder will rangefrom about 10 μg/day to about 200 μg/day.

When administered by inhalation, the compounds of this inventiontypically have the effect of providing bronchodilation. Accordingly, inanother of its method aspects, this invention is directed to a method ofproviding bronchodilation in a patient, the method comprisingadministering to a patient requiring bronchodilation a therapeuticallyeffective amount of a compound of formula I or a pharmaceuticallyacceptable salt or solvate or stereoisomer thereof. Generally, the dosefor providing bronchodilation will range from about 10 μg/day to about200 μg/day.

In one embodiment, this invention is directed to a method of treatingchronic obstructive pulmonary disease or asthma, the method comprisingadministering to a patient in need of treatment a therapeuticallyeffective amount of a compound of formula I or a pharmaceuticallyacceptable salt or solvate or stereoisomer thereof. When used to treat aCOPD or asthma, the compounds of this invention will typically beadministered by inhalation in multiple doses per day or in a singledaily dose. Generally, the dose for treating COPD or asthma will rangefrom about 10 μg/day to about 200 μg/day.

As used herein, COPD includes chronic obstructive bronchitis andemphysema (see, for example, Barnes, Chronic Obstructive PulmonaryDisease, N Engl J Med 2000: 343:269-78).

When used to treat a pulmonary disorder, the compounds of this inventionare optionally administered in combination with other therapeuticagents. In particular, by combining the compounds of this invention witha steroidal anti-inflammatory agent (e.g. a corticosteroid), thepharmaceutical compositions of this invention can provide tripletherapy, i.e., β₂ adrenergic receptor agonist, muscarinic receptorantagonist and anti-inflammatory activity, using only two activecomponents. Since pharmaceutical compositions containing two activecomponents are typically easier to formulate compared to compositionscontaining three active components, such two component compositionsprovide a significant advantage over compositions containing threeactive components. Accordingly, in a particular embodiment, thepharmaceutical compositions and methods of this invention furthercomprise a therapeutically effective amount of a steroidalanti-inflammatory agent.

Compounds of this invention exhibit both muscarinic receptor antagonistand β₂ adrenergic receptor agonist activity. Accordingly, among otherproperties, compounds of particular interest are those that demonstratean inhibitory constant K_(i) value for binding at the M₃ muscarinicreceptor and an EC₅₀ value for β₂ adrenergic receptor agonist activityof less than about 100 nM; particularly less than 10 nM. Among thesecompounds, compounds of special interest include those having muscarinicactivity, expressed in terms of the inhibitory constant K_(i) forbinding at the M₃ muscarinic receptor, that is about equal to thecompound's β₂ adrenergic agonist activity, expressed in terms of thehalf maximal effective concentration EC₅₀, as determined in the in vitroassays described herein, or in similar assays. For example, compounds ofparticular interest are those having a ratio of the inhibitory constantK_(i) for the M₃ muscarinic receptor to the EC₅₀ for the β₂ adrenergicreceptor ranging from about 30:1 to about 1:30; including about 20:1 toabout 1:20; such as about 10:1 to about 1:10.

In one of its method aspects, the present invention also provides amethod for treating a pulmonary disorder, the method comprisingadministering to a patient in need of treatment a therapeuticallyeffective amount of a compound having both muscarinic receptorantagonist and β₂ adrenergic receptor agonist activity. In a particularembodiment of this method, the compound administered has an inhibitoryconstant K_(i) for the M₃ muscarinic receptor that is less than about100 nM and a half maximal effective concentration EC₅₀ for agonism atthe β₂ adrenergic receptor that is less than about 100 nM. In anotherembodiment, the method for treating a pulmonary disorder comprisesadministering a therapeutically effective amount of a compound for whichthe ratio of the inhibitory constant K_(i) for the M₃ muscarinicreceptor to the EC₅₀ for agonism of the β₂ adrenergic receptor isbetween about 30:1 and about 1:30.

Since compounds of this invention possess both β₂ adrenergic agonistactivity and muscarinic receptor antagonist activity, such compounds arealso useful as research tools for investigating or studying biologicalsystems or samples having β₂ adrenergic receptors or muscarinicreceptors, or for discovering new compounds having both β₂ adrenergicagonist activity and muscarinic receptor antagonist activity. Suchbiological systems or samples may comprise β₂ adrenergic receptorsand/or muscarinic receptors. Any suitable biological system or samplehaving β₂ adrenergic and/or muscarinic receptors may be employed in suchstudies which may be conducted either in vitro or in vivo.Representative biological systems or samples suitable for such studiesinclude, but are not limited to, cells, cellular extracts, plasmamembranes, tissue samples, mammals (such as mice, rats, guinea pigs,rabbits, dogs, pigs, etc.), and the like.

In this embodiment, a biological system or sample comprising a β₂adrenergic receptor or a muscarinic receptor is contacted with a β₂adrenergic receptor-agonizing or muscarinic receptor-antagonizing amountof a compound of this invention. The effects are then determined usingconventional procedures and equipment, such as radioligand bindingassays and functional assays. Such functional assays includeligand-mediated changes in intracellular cyclic adenosine monophosphate(cAMP), ligand-mediated changes in activity of the enzyme adenylylcyclase (which synthesizes cAMP), ligand-mediated changes inincorporation of guanosine 5′-O-(-thio)triphosphate ([³⁵S]GTP S) intoisolated membranes via receptor catalyzed exchange of [³⁵S]GTP S forGDP, ligand-mediated changes in free intracellular calcium ions(measured, for example, with a fluorescence-linked imaging plate readeror FLIPR® from Molecular Devices, Inc.). A compound of this inventionwill agonize or cause activation of a β₂ adrenergic receptor andantagonize or decrease the activation of muscarinic receptors in any ofthe functional assays listed above, or assays of a similar nature. Theamount of compound used in these studies will typically range from about0.1 nanomolar to about 100 nanomolar.

Additionally, the compounds of this invention can be used as researchtools for discovering new compounds that have both a β₂ adrenergicreceptor agonist and muscarinic receptor antagonist activity. In thisembodiment, a β₂ adrenergic receptor and muscarinic receptor bindingdata (for example, as determined by in vitro radioligand displacementassays) for a test compound or a group of test compounds is compared tothe β₂ adrenergic receptor and muscarinic receptor binding data for acompound of this invention to identify those test compounds that haveabout equal or superior β₂ adrenergic receptor and/or muscarinicreceptor binding, if any. This aspect of the invention includes, asseparate embodiments, both the generation of comparison data (using theappropriate assays) and the analysis of the test data to identify testcompounds of interest.

In some cases, compounds of this invention may possess either weakmuscarinic receptor antagonist activity or weak β₂ adrenergic receptoragonist activity. In these cases, those of ordinary skill in the artwill recognize that such compounds still have utility as primarilyeither a β₂ adrenergic receptor agonist or a muscarinic receptorantagonist, respectively.

The properties and utility of the compounds of this invention can bedemonstrated using various in vitro and in vivo assays well-known tothose skilled in the art. For example, representative assays aredescribed in further detail in the following Examples.

EXAMPLES

The following Preparations and Examples are provided to illustratespecific embodiments of this invention. These specific embodiments,however, are not intended to limit the scope of this invention in anyway unless specifically indicated.

The following abbreviations have the following meanings unless otherwiseindicated and any other abbreviations used herein and not defined havetheir standard meaning:

AC adenylyl cyclase Ach acetylcholine ATCC American Type CultureCollection BSA bovine serum albumin cAMP 3′-5′ cyclic adenosinemonophosphate CHO Chinese hamster ovary cM₅ cloned chimpanzee M₅receptor DCM dichloromethane (i.e., methylene chloride) DIPEAN,N-diisopropylethylamine dPBS Dulbecco's phosphate buffered saline DMEMDulbecco's Modified Eagle's Medium DMSO dimethyl sulfoxide EDTAethylenediaminetetraacetic acid Emax maximal efficacy EtOAc ethylacetate EtOH ethanol FBS fetal bovine serum FLIPR fluorometric imagingplate reader Gly glycine HATUO-(7-azabenzotriazol-1-yl-N,N,N′,N′-tetramethyluroniumhexafluorophosphate HBSS Hank's buffered salt solution HEK humanembryonic kidney cells HEPES4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid hM₁ cloned human M₁receptor hM₂ cloned human M₂ receptor hM₃ cloned human M₃ receptor hM₄cloned human M₄ receptor hM₅ cloned human M₅ receptor HPLChigh-performance liquid chromatography IBMX 3-isobutyl-1-methylxanthine% Eff % efficacy PBS phosphate buffered saline PyBOPbenzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate rpmrotations per minute TFA trifluoroacetic acid THF tetrahydrofuran Tristris(hydroxymethyl)aminomethane

Unless noted otherwise, reagents, starting materials and solvents werepurchased from commercial suppliers (such as Aldrich, Fluka, Sigma andthe like) and were used without further purification.

In the examples described below, HPLC analysis was conducted using thefollowing conditions where indicated:

HPLC Method A:

Column: YMC ODSA 5μ C18 4.6 × 50 mm Detector Wavelength: 220 nm ColumnTemperature: 35° C. Flow Rate: 4.0 mL/min Solvent System: A = 10%methanol, 90% water, 0.1% TFA B = 90% methanol, 10% water, 0.1% TFAInjection Volume: 5 μL Run Time: 5 min Gradient: 0-100% B in AHPLC Method B:

Column: Inertsil OCD-2 C18 Detector Wavelength: 254 nm ColumnTemperature: 35° C. Flow Rate: 1.0 mL/min Solvent System: A = 5%methanol, 95% water, 0.1% TFA B = 95% methanol, 5% water, 0.1% TFAInjection Volume: 5 μL Run Time: 15 min Gradient: 0-100% B in A

Liquid chromatography mass spectrometry (LCMS) data were obtained withan Applied Biosystems (Foster City, Calif.) model API-150EX instrument.

Small-scale purification was conducted using an API 150EX PrepWorkstation system from Applied Biosystems. The mobile phase was A:water+0.05% v/v TFA; and B: acetonitrile+0.05% v/v TFA. For arrays(typically about 3 to 50 mg recovered sample size) the followingconditions were used: 20 mL/min flow rate; 15 min gradients and a 20mm×50 mm Prism RP column with 5 micron particles (ThermoHypersil-Keystone, Bellefonte, Pa.). For larger scale purifications(typically greater than 100 mg crude sample), the following conditionswere used: 60 mL/min flow rate; 30 min gradients and a 41.4 mm×250 mmMicrosorb BDS column with 10 micron particles (Varian, Palo Alto,Calif.).

The specific rotation for chiral compounds (indicated as [α]²⁰D) wasmeasured using a Jasco Polarimeter (Model P-1010) with a tungstenhalogen light source and a 589 nm filter at 20° C. Samples of testcompounds were typically measured at 1 mg/mL water.

Preparation 1 2,2-Diphenyl-2-(S)-pyrrolidin-3-ylacetamide

(a) Toluene-4-sulfonic Acid (S)-1-Benzylpyrrolidin-3-yl Ester

To a stirred solution of (S)-1-benzyl-3-pyrrolidinol (44.3 g, 0.25 mol)and 1,4-diazabicyclo[2.2.2]octane (33.7 g, 0.3 mol) in 250 mL oftert-butyl methyl ether under an atmosphere of nitrogen at 0° C., wasadded p-toluenesulfonyl chloride (52.4 g, 0.275 mol) portionwise over 20min. The reaction mixture was stirred at 0° C. for 1 h. The ice bath wasthen removed and the mixture was stirred at ambient temperatureovernight (20±5 h). Ethyl acetate (100 mL) was then added, followed bysaturated aqueous sodium bicarbonate solution (250 mL). The resultingmixture was stirred at ambient temperature for 1 h. The layers wereseparated and the organic layer was washed with saturated aqueous sodiumbicarbonate solution (250 mL); saturated aqueous ammonium chloridesolution (250 mL); saturated aqueous sodium chloride solution (250 mL);and then dried over sodium sulfate (80 g). The sodium sulfate wasfiltered off and washed with ethyl acetate (20 mL) and the solvent wasremoved in vacuo to give 78.2 g of the title intermediate (94% yield;95% purity by HPLC Method A).

(b) ((S)-1-Benzylpyrrolidin-3-yl)diphenylacetonitrile

To a stirred solution of diphenylacetonitrile (12.18 g, 61.8 mmol) inanhydrous THF (120 mL) at 0° C., potassium tert-butoxide (10.60 g, 94.6mmol) was added over 5 min. The reaction mixture was stirred at 0° C.for 1 h. The product of step (a) above (20.48 g, 61.3 mmol) was thenadded to the reaction mixture at 0° C. in one portion. The cold bath wasremoved and the reaction mixture was stirred for 5-10 min, at which timethe reaction mixture had become a homogeneous solution. The reactionmixture was then heated at 40° C. overnight (20±5 h). The reactionmixture was allowed to cool to ambient temperature, and then water (150mL) was added. Most of the THF was then removed in vacuo and isopropylacetate (200 mL) was added. The layers were separated and the organiclayer was washed with saturated aqueous ammonium chloride solution (150mL) and saturated aqueous sodium chloride solution (150 mL); and thendried over sodium sulfate (50 g). The sodium sulfate was filtered offand washed with isopropyl acetate (20 mL). The solvent was removed invacuo to give 23.88 g of the title intermediate as an oil (>99% yield,75% purity by HPLC Method A, containing mainly an excess ofdiphenylacetonitrile starting material).

(c) Diphenyl-(S)-pyrrolidin-3-ylacetonitrile

The product of step (b) above was dissolved in isopropyl acetate (ca. 1g/10 mL) and the solution was mixed with an equal volume of 1N aqueoushydrochloric acid. The resulting layers were separated and the aqueouslayer was extracted with an equal volume of isopropyl acetate. Theorganic layers were combined, dried over sodium sulfate and filtered.The solvent was removed in vacuo to afford((S)-1-benzylpyrrolidin-3-yl)diphenylacetonitrile hydrochloride as afoamy solid. (Note: This hydrochloride salt can also be prepared duringthe work-up of Step (b)).

To a stirred solution of((S)-1-benzylpyrrolidin-3-yl)diphenylacetonitrile hydrochloride (8.55 g,21.98 mmol) in methanol (44 mL) was added palladium on carbon (1.71 g)and ammonium formate (6.93 g, 109.9 mmol). The reaction mixture washeated to 50° C. with stirring for 3 h. The reaction was cooled toambient temperature and water (20 mL) was added. The resulting mixturewas filtered through a pad of Celite, washing with methanol (20 mL). Thefiltrate was collected and most of the methanol was removed in vacuo.The residue was mixed with isopropyl acetate (100 mL) and 10% aqueoussodium carbonate (50 mL). The resulting layers were separated and theaqueous layer was extracted with isopropyl acetate (50 mL). The organiclayers were combined and dried over sodium sulfate (20 g). The sodiumsulfate was filtered off and washed with isopropyl acetate (20 mL). Thesolvent was removed in vacuo to afford 5.75 g of the title intermediateas an oil (99.7% yield, 71% purity by HPLC).

(d) 2,2-Diphenyl-2-(S)-pyrrolidin-3-ylacetamide

A 200 mL flask with a magnetic stir bar and a nitrogen inlet was chargedwith the product from step (c) (2.51 g) and 80% sulfuric acid (19.2 mL;pre-prepared with 16 mL of 96% sulfuric acid and 3.2 mL of water). Thereaction mixture was then heated at 90° C. for 24 h or until startingmaterial was consumed as indicated by HPLC. The reaction mixture wasallowed to cool to ambient temperature and then poured onto ice (ca. 50mL by volume). A 50% aqueous sodium hydroxide solution was added slowlyto the mixture with stirring over an ice bath until the pH was about 12.Dichloromethane (200 mL) was added and mixed with the aqueous solution,at which time sodium sulfate precipitated out and was filtered off. Thefiltrate was collected and the layers were separated. The aqueous layerwas extracted with dichloromethane (100 mL) and the organic layers werecombined dried with over sodium sulfate (5 g). The sodium sulfate wasfiltered off and washed with dichloromethane (10 mL). The solvent wasremoved in vacuo to give the crude product as a light yellow foamy solid(ca. 2.2 g, 86% purity by HPLC).

The crude product was dissolved in ethanol (18 mL) with stirring. Tothis solution was added a warm solution of L-tartaric acid (1.8 g) inethanol (14 mL) and the resulting mixture was stirred overnight (15±5h). The resulting precipitate was isolated by filtration to give anoff-white solid (ca. 3.2 g, >95% purity by HPLC). Methanol (15 mL) wasadded to this solid and the resulting slurry was stirred at 70° C.overnight (15 h). The slurry was allowed to cool to ambient temperatureand a solid (˜2.6 g, >99% purity by HPLC) was obtained after filtration.To this solid was added ethyl acetate (30 mL) and 1 N aqueous sodiumhydroxide (25 mL). This mixture was stirred until two distinct layersformed and then the layers were separated and the aqueous layer wasextracted with ethyl acetate (20 mL). The organic layers were combinedand dried over sodium sulfate (10 g). The sodium sulfate was removed byfiltration and the solvent was evaporated in vacuo to afford 1.55 g ofthe title intermediate as a solid (58% yield; >99% purity by HPLC MethodB).

Preparation 22-[(S)-1-(9-Hydroxynonyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

9-Bromo-1-nonanol (3.50 g, 15.7 mmol) in acetonitrile (8 mL) was addedto a stirred solution of the product of Preparation 1 (4.0 g, 14.3 mmol)and triethylamine (5.98 mL, 42.9 mmol) in acetonitrile (135 mL) at 40°C. The reaction mixture was heated at 50° C. for 24 h and then cooled toambient temperature. The solvent was then removed under reducedpressure. The crude residue was dissolved in DCM (100 mL) and theorganic phase was washed with saturated aqueous sodium bicarbonate (50mL), dried over magnesium sulfate, filtered and concentrated underreduced pressure. The crude product was purified by flash chromatography(5-10% MeOH in DCM containing 0.5% ammonium hydroxide) to give the titlecompound as a solid (3.77 g, 8.93 mmol, 62% yield).

Preparation 32-[(S)-1-(9-Oxononyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

Dimethyl sulfoxide (6.34 mL, 89.3 mmol) and diisopropylethylamine (7.79mL, 44.7 mmol) were added to a stirred solution of the product ofPreparation 2 (3.77 g, 8.93 mmol) in dichloromethane (89.3 mL) at 0° C.The reaction mixture was stirred at 0° C. for 15 min and then sulfurtrioxide pyridine complex (7.11 g, 44.7 mmol) was added. This mixturewas stirred for 1 h at 0° C. and then water (50 mL) was added. Theorganic layer was separated and washed with saturated aqueous copper(II) sulfate solution (3×30 mL), brine (30 mL), dried over magnesiumsulfate, filtered and concentrated under reduced pressure to provide thetitle compound as a solid (˜100% yield).

Preparation 42-[(S)-1-(9-Benzylaminononyl)pyrrolidin-3-yl]-2,2-diphenylacetanide

The product of Preparation 3 (420 mg, 1 mmol) and benzyl amine (328 μL,3 mmol) in methanol (5 mL) were stirred at ambient temperature for 12 hand then sodium cyanoborohydride (94 mg, 1.5 mmol) in methanol (5 mL)was added. This mixture was stirred for 24 h and then the solvent wasremoved under reduced pressure. The residue was dissolved in DCM (15 mL)and the organic phase was washed with saturated aqueous sodiumbicarbonate solution (8 mL), dried over magnesium sulfate, filtered andconcentrated under reduced pressure. The residue was purified bypreparative HPLC to give the title compound as the ditrifluoroaceticacid salt (340 mg, 46% yield).

Preparation 5 8-Benzyloxy-5-(2-bromoacetyl)-1H-quinolin-2-one

(a) 8-Acetoxy-1H-quinolin-2-one

8-Hydroxyquinoline-N-oxide (160.0 g, 1.0 mol), commercially-availablefrom Aldrich, Milwaukee, Wis., and acetic anhydride (800 mL, 8.4 mol)were heated at 100° C. for 3 h and then cooled in ice. The product wascollected on a Buchner funnel, washed with acetic anhydride (2×100 mL)and dried under reduced pressure to give 8-acetoxy-1H-quinolin-2-one(144 g) as a solid.

(b) 5-Acetyl-8-hydroxy-1H-quinolin-2-one

A slurry of aluminum chloride (85.7 g, 640 mmol) in 1,2-dichloroethane(280 mL) was cooled in ice, and the product from step (a) (56.8 g, 280mmol) was added. The mixture was warmed to room temperature and thenheated at 85° C. After 30 min, acetyl chloride (1.5 mL, 21 mmol) wasadded and the mixture was heated an additional 60 min. The reactionmixture was then cooled and added to 1N hydrochloric acid (3 L) at 0° C.with good stirring. After stirring for 2 h, the solids were collected ona Buchner funnel, washed with water (3×250 mL) and dried under reducedpressure. The crude product isolated from several batches (135 g) wascombined and triturated with dichloromethane (4 L) for 6 h. Theresulting solid was collected on a Buchner funnel and dried underreduced pressure to give the title compound (121 g).

(c) 5-Acetyl-8-benzyloxy-1H-quinolin-2-one

To the product from step (b) (37.7 g, 186 mmol) was addedN,N-dimethylformamide (200 mL) and potassium carbonate (34.5 g, 250mmol) followed by benzyl bromide (31.8 g, 186 mmol). The mixture wasstirred at room temperature for 2.25 hour and then poured into saturatedsodium chloride (3.5 L) at 0° C. and stirred for 1 hour. The product wascollected and dried on a Buchner funnel for 1 hour, and the resultingsolids were dissolved in dichloromethane (2 L) and this mixture wasdried over sodium sulfate. The solution was filtered through a pad ofCelite which was then washed with dichloromethane (5×200 mL). Thecombined filtrate was then concentrated to dryness and the resultingsolids were triturated with ether (500 mL) for 2 h. The product wascollected on a Buchner funnel, washed with ether (2×250 mL) and driedunder reduced pressure to give the title compound (44 g) as a powder.

(d) 8-Benzyloxy-5-(2-bromoacetyl)-1H-quinolin-2-one

The product from step (c) (20.0 g, 68.2 mmol) was dissolved indichloromethane (200 mL) and cooled to 0° C. Boron trifluoride diethyletherate (10.4 mL, 82.0 mmol) was added via syringe and the mixture waswarmed to room temperature to give a thick suspension. The suspensionwas heated at 45° C. (oil bath) and a solution of bromine (11.5 g, 72.0mmol) in dichloromethane (100 mL) was added over 40 min. The mixture waskept at 45° C. for an additional 15 min and then cooled to roomtemperature. The mixture was concentrated under reduced pressure andthen triturated with 10% aqueous sodium carbonate (200 mL) for 1 hour.The solids were collected on a Buchner funnel, washed with water (4×100mL) and dried under reduced pressure. The product of two runs wascombined for purification. The crude product (52 g) was triturated with50% methanol in chloroform (500 mL) for 1 hour. The product wascollected on a Buchner funnel and washed with 50% methanol in chloroform(2×50 mL) and methanol (2×50 mL). The solid was dried under reducedpressure to give the title compound (34.1 g) as a powder.

Preparation 62-[(S)-1-(9-{Benzyl-[2-(8-benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-oxoethyl]-amino}nonyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

To the product of Preparation 4 (256 mg, 0.5 mmol) and potassiumcarbonate (207 mg, 1.5 mmol) in N,N-dimethylformamide (2.5 mL) was addedthe product of Preparation 5 (372 mg, 1.0 mmol). The reaction mixturewas heated at 45° C. for 24 h and then filtering through a cotton plug.The solvent was removed under reduced pressure to yield the titlecompound, which was used without further purification.

Preparation 72-[(S)-1-(9-{Benzyl-[2-(8-benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethyl]-amino}nonyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 6 (401 mg, 0.5 mmol)in ethanol (2.5 mL), was added sodium borohydride (57 mg, 1.5 mmol) andN,N-dimethylformamide (2.5 mL). The reaction mixture was stirred for 6 hat room temperature and then the solvent was removed under reducedpressure. Ethyl acetate (10 mL) was added to the crude residue and theorganic layer was washed with 10% potassium carbonate solution (5 mL),dried over magnesium sulfate, filtered and concentrated under reducedpressure. The residue was purified by flash chromatography (5% MeOH:DCM)to give the title compound (202 mg, 0.25 mmol, 50% over 2 steps).

Example 12-((S)-1-{9-[2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]-nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

Palladium (40 mg, 10 wt. % (dry basis) on activated carbon) was added toa stirred solution of the product of Preparation 7 (202 mg, 0.25 mmol)in acetic acid (2.5 mL) and the resulting mixture was placed under ahydrogen atmosphere and stirred overnight. The reaction mixture was thenfiltered and the solvent was removed under reduced pressure. The residuewas purified by preparative HPLC to afford the title compound as theditrifluoroacetate salt (20 mg). HPLC (10-70) R_(t)=2.84.

Preparation 8 N,N-(Di-tert-butoxycarbonyl)-9-bromononylamine

A solution of di-tert-butoxycarbonylamine (3.15 g, 14.5 mmol) inN,N-dimethylformamide (0.28 mL) was cooled to 0° C. for about 10 min.Sodium hydride, 60% in mineral oil (0.58 g, 14.5 mmol) was added and thereaction mixture was stirred at 0° C. for 10 min. The reaction mixturewas removed from the ice bath and allowed to warm to room temperaturefor about 30 min. The reaction mixture was then cooled back down to 0°C. and a solution of 1,9-dibromononane (2.46 mL, 12.1 mmol) indimethylformamide (100 mL) was added. The reaction mixture was stirredovernight at room temperature. After 24 h, MS analysis showed that thereaction was completed. The reaction mixture was concentrated to drynessand diluted with ethyl acetate (100 mL). The organic layer was washedwith saturated sodium bicarbonate (2×100 mL), brine (100 mL), dried(magnesium sulfate) and concentrated under reduced pressure to yield thecrude product, which was purified by chromatography on silica gel using5% ethyl acetate in hexanes to afford the title compound. MS m/z: [M+H⁺]calcd for C₁₉H₃₆N₁O₄Br 423.18. found 423.

Preparation 92-[(S)-1-(9-Di-tert-butoxycarbonylaminononyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

A mixture of 1:1 acetonitrile and N,N-dimethylformamide (50 mL) is addedto the products of Preparation 1 (10.1 mmol) and Preparation 8 (5.1 g,12.2 mmol) and triethylamine (1.42 mL, 10.1 mmol). The reaction mixtureis stirred at ambient temperature for 24 h. The reaction mixture is thenconcentrated and diluted with ethyl acetate (50 mL). The organic layeris washed with saturated sodium bicarbonate (2×50 mL) and brine (50 mL).The organic phase is then dried over magnesium sulfate and concentrated.The residue is purified by chromatography to provide the title compound.

Preparation 102-[(S)-1-(9-Aminononyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

Trifluoroacetic acid (11 mL) is added to a solution of the product ofPreparation 9 (11.3 mmol) in dichloromethane (56 mL). After 2 h, thereaction mixture is concentrated to dryness and diluted with ethylacetate (75 mL). Sodium hydroxide (1N) is then added until the pH of themixture reaches 14. The organic phase is then collected and washed withsaturated sodium bicarbonate (2×50 mL) and brine (50 mL). The organicphase is then dried over magnesium sulfate and concentrated to providethe title compound.

Preparation 118-Benzyloxy-5-[(R)-2-bromo-1-(tert-butyldimethylsilanyloxy)ethyl]-1H-quinolin-2-one

(a) 8-Benzyloxy-5-((R)-2-bromo-1-hydroxyethyl)-1H-quinolin-2-one

(R)-(+)-α,α-Diphenylprolinol (30.0 g, 117 mmol) and trimethylboroxine(11.1 mL, 78 mmol) were combined in toluene (300 mL) and stirred at roomtemperature for 30 min. The mixture was placed in a 150° C. oil bath andliquid was distilled off. Toluene was added in 20 mL aliquots anddistillation was continued for 4 h. A total of 300 mL toluene was added.The mixture was then cooled to room temperature. A 500 μL aliquot wasevaporated to dryness and weighed (246 mg) to determine that theconcentration of catalyst was 1.8 M.

8-Benzyloxy 5-(2-bromoacetyl)-1H-quinolin-2-one (90.0 g, 243 mmol) wasplaced under nitrogen and tetrahydrofuran (900 mL) was added followed bythe catalyst described above (1.8 M in toluene, 15 mL, 27 mmol). Thesuspension was cooled to −10±5° C. in an ice/isopropanol bath. Borane(1.0 M in THF, 294 mL, 294 mmol) was added over 4 h. The reaction wasthen stirred an additional 45 min at −10° C. and then methanol (250 mL)was added slowly. The mixture was concentrated under vacuum and theresidue was dissolved in boiling acetonitrile (1.3 L), filtered whilehot and then cooled to room temperature. The crystals were filtered,washed with acetonitrile and dried under vacuum to give the titlecompound (72.5 g, 196 mmol, 81% yield, 95% ee, 95% pure by HPLC).

(b)8-Benzyloxy-5-[(R)-2-bromo-1-(tert-butyldimethylsilanyloxy)ethyl]-1H-quinolin-2-one

To the product of step (b) (70.2 g, 189 mmol) was addedN,N-dimethylformamide (260 mL) and this mixture was cooled in an icebath under nitrogen. 2,6-Lutidine (40.3 g, 376 mmol) was added over 5min and then tert-butyldimethylsilyl trifluoromethanesulfonate (99.8 g,378 mmol) was added slowly while maintaining the temperature below 20°C. The mixture was allowed to warm to room temperature for 45 min.Methanol (45 mL) was added to the mixture dropwise over 10 min and themixture was partitioned between ethyl acetate/cyclohexane (1:1, 500 mL)and water/brine (1:1, 500 mL). The organics were washed twice more withwater/brine (1:1, 500 mL each). The combined organics were evaporatedunder reduced pressure to give a light yellow oil. Two separate portionsof cyclohexane (400 mL) were added to the oil and distillation continueduntil a thick white slurry was formed. Cyclohexane (300 mL) was added tothe slurry and the resulting white crystals were filtered, washed withcyclohexane (300 mL) and dried under reduced pressure to give the titlecompound (75.4 g, 151 mmol, 80% yield, 98.6% ee).

Preparation 122-((S)-1-{9-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyl-dimethylsilanyloxy)ethylamino]nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

The product of Preparation 11 (3.9 g, 8.17 mmol) is added to a solutionof the product of Preparation 10 (11.4 mmol) in THF (20 mL), followed bysodium bicarbonate (2.0 g, 24.5 mmol) and sodium iodide (1.8 g, 12.2mmol). The reaction mixture is heated to 80° C. for 72 h. The reactionmixture is then cooled, diluted with dichloromethane (20 mL) and theorganic phase is washed with saturated sodium bicarbonate (2×50 mL) andbrine (50 mL). The organic phase is then dried (magnesium sulfate) andconcentrated to give a crude product. The crude product is purified bychromatography on silica gel to provide the title compound.

Preparation 132-((S)-1-{9-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxy-ethylamino]nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

Triethylamine hydrogen fluoride (376 μL, 2.3 mmol) is added to asolution of the product of Preparation 12 (1.5 mmol) in THF (8 mL) andthe reaction mixture is stirred at ambient temperature. After 5 h, thereaction mixture is quenched with 1N NaOH until the pH is 14 and thendiluted with ethyl acetate (20 mL) and washed with 1N NaOH (20 mL) andbrine (20 mL). The organic phase is then separated, dried over magnesiumsulfate, and concentrated to yield the title compound.

Example 22-((S)-1-{9-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

A solution of the product of Preparation 13 (1.5 mmol) is flushed withnitrogen and palladium on carbon (10%, 110 mg) is added. The reactionmixture is stirred under hydrogen at balloon pressure. After 9 h, thereaction mixture is filtered and concentrated. The residue is purifiedby HPLC to afford the title compound as the ditrifluoroacetate salt.

Preparation 143-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]propionic acid MethylEster

Methyl 3-bromopropionate (553 μL, 5.07 mmol) is added to a stirredsolution of the product of Preparation 1 (3.38 mmol) and DIPEA (1.76 mL,10.1 mmol) in acetonitrile (34 mL) at 50° C. and the reaction mixture isheated at 50° C. overnight. The solvent is then removed under reducedpressure and the residue is dissolved in dichloromethane (30 mL). Theresulting solution is washed with saturated aqueous sodium bicarbonatesolution (10 mL), dried (magnesium sulfate) and the solvent is removedunder reduced pressure. The crude residue is purified by columnchromatography to give the title compound.

Preparation 153-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]propionic Acid

A stirred solution of the product of Preparation 14 (2.37 mmol) andlithium hydroxide (171 mg, 7.11 mmol) in 50% THF/H₂O (24 mL) is heatedat 30° C. overnight, and then acidified with concentrated hydrochloricacid and lyophilized to give the title compound.

Preparation 16{5-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]pentyl}carbamicAcid tert-Butyl Ester

The product of Preparation 11 (600 mg, 1.23 mmol) andN-tert-butoxycarbonyl-1,5-diaminopentane (622 mg, 3.07 mmol) weredissolved in dimethyl sulfoxide (1.23 mL) and heated to 105° C. for 6 h.The reaction mixture was then cooled and diluted with ethyl acetate (10mL) and washed with saturated aqueous sodium bicarbonate solution (4mL). The organic phase was dried (magnesium sulfate) and the solvent wasremoved under reduced pressure. The crude residue was purified by columnchromatography (5-10% methanol/dichloromethane) to give the titlecompound (˜100% yield).

Preparation 175-[(R)-2-(5-Aminopentylamino)-1-(tert-butyldimethylsilanyloxy)ethyl]-8-benzyloxy-1H-quinolin-2-one

A solution of the product of Preparation 16 (800 mg, 1.31 mmol) intrifluoroacetic acid/dichloromethane (25%, 12 mL) was stirred at ambienttemperature for 1 hour. The solvent was then removed under reducedpressure and the crude residue was dissolved in dichloromethane (15 mL)and washed with 1N sodium hydroxide (8 mL). The organic phase wasseparated, dried (magnesium sulfate) and the solvent was removed underreduced pressure to give the title compound (509 mg, 81% yield over 2steps).

Preparation 18N-{5-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]pentyl}-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]-propionamide

To the product of Preparation 15 (1.13 mmol) and HATU (430 mg, 1.13mmol) is added the product of Preparation 17 (458 mg, 0.90 mmol) in DMF(1.8 mL), followed by DIPEA (204 μL, 1.17 mmol). The reaction mixture isstirred at 50° C. for 12 h, and then the solvent is removed underreduced pressure. The crude residue is dissolved in dichloromethane (10mL). The resulting solution is washed with saturated aqueous sodiumbicarbonate solution (4 mL), dried (magnesium sulfate) and the solventis removed under reduced pressure. The crude residue is purified bycolumn chromatography to give the title compound.

Preparation 19N-{5-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethylamino]-pentyl}-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]propionamide

To a stirred solution of the product of Preparation 18 (0.28 mmol) indichloromethane (2.8 mL) is added triethylamine trihydrofluoride (91 μL,0.56 mmol). The reaction mixture is stirred for 10 h, and then dilutedwith dichloromethane (10 mL). The resulting solution is then washed withsaturated aqueous sodium bicarbonate solution (5 mL), and then theorganic phase is dried (magnesium sulfate) and the solvent is removedunder reduced pressure to give the title compound.

Example 33-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-N-{5-[(R)-2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]pentyl}propionamide

To a stirred solution of the product of Preparation 19 (0.28 mmol) inethanol (2.8 mL) is added palladium (10 wt. % (dry basis) on activatedcarbon) (81 mg) and the reaction mixture is placed under an atmosphereof hydrogen and stirred overnight. The reaction mixture is then filteredand solvent is removed under reduced pressure. The crude residue ispurified by preparative HPLC to give the title compound as theditrifluoroacetate salt.

Preparation 20 2-(N-Benzyloxycarbonyl-N-methylamino)ethanal

(a) 2-(N-Benzyloxycarbonyl-N-methylamino)ethanol

Benzyl chloroformate (19 g, 111.1 mmol) in THF (20 mL) was addeddropwise over 15 min to a stirred solution of 2-(methylamino)ethanol (10g, 133.3 mmol) in THF (100 mL) and aqueous sodium carbonate (100 mL) at0° C. The reaction mixture was stirred at 0° C. for 12 h and thenextracted with EtOAc (2×200 mL). The organic layer was washed withaqueous sodium carbonate (200 mL) and dried (potassium carbonate) andsolvent was removed under reduced pressure to give the title compound(22.5 g, 97% yield).

(b) 2-(N-Benzyloxycarbonyl-N-methylamino)ethanal

DMSO (71 mL, 1 mol) and DIPEA (87.1 mL, 0.5 mol) were added to a stirredsolution of the product of step (a) (20.9 g, 0.1 mol) in dichloromethane(200 mL) at −10° C. The reaction mixture was stirred at −10° C. for 15min and then sulfur trioxide pyridine complex (79.6 g, 0.5 mol) wasadded and the resulting mixture was stirred for 1 hour. The reactionmixture was quenched with addition of 1 M hydrochloric acid (200 mL).The organic layer was separated and washed with saturated aqueous sodiumbicarbonate (100 mL), brine (100 mL), dried (potassium carbonate) andsolvent removed under reduced pressure to give the title compound (20.7g, ˜100% yield).

Preparation 212-[(S)-1-(2-Methylaminoethyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 20 (20.7 g, 100mmol) and the product of Preparation 1 (84.7 mmol) in MeOH (200 mL) isadded sodium triacetoxyborohydride (21.2 g, 100 mmol). The reactionmixture is stirred for 12 h at ambient temperature and then it isquenched with 2M hydrochloric acid and the solvent is removed underreduced pressure. The residue is dissolved in ethyl acetate (200 mL) andwashed with saturated aqueous sodium bicarbonate solution (100 mL) andbrine (50 mL), and then dried (magnesium sulfate) and the solvent isremoved under reduced pressure. The crude residue is purified by columnchromatography (50-90% EtOAc/hexanes) to give2-[(S)-1-(2-benzyloxycarbonylmethylaminoethyl)pyrrolidin-3-yl]-2,2-diphenylacetamide.This material is dissolved in methanol (100 mL) and palladium (10 wt. %(dry basis) on activated carbon) (5 g) is added. The reaction mixture isstirred under hydrogen (30 psi) for 12 h and then filtered throughCelite, which is washed with methanol, and solvent is evaporated to givethe title compound.

Preparation 22 6-Bromohexanoic Acid{2-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]ethyl}-methylamide

6-Bromohexanoyl chloride (3.23 mL, 21.1 mmol) is added to a stirredsolution of the product of Preparation 21 (17.6 mmol) and DIPEA (6.13mL, 35.2 mmol) in dichloroethane (170 mL). The reaction mixture isstirred for 1 hour and it is then diluted with EtOAc (250 mL) and washedwith saturated aqueous sodium bicarbonate solution (2×200 mL) and brine(200 mL), and then dried (magnesium sulfate). The solvent is removedunder reduced pressure to give the title compound.

Preparation 238-Benzyloxy-5-[(R)-2-(N-benzylamino)-1-(tert-butyldimethylsilanyloxy)ethyl]1H-quinolin-2-one

A stirred solution of the product of Preparation 11 (1.00 g, 2.05 mmol)and benzylamine (493 μL, 4.51 mmol) in DMSO (1.7 mL) was heated at 105°C. for 4 h. The reaction mixture was allowed to cool and was thendiluted with EtOAc (10 mL) and the organic layer was washed withsaturated aqueous ammonium chloride solution (5 mL) and 1N sodiumhydroxide (5 mL), dried (MgSO₄) and solvent removed under reducedpressure. The crude residue was purified by column chromatography (50%EtOAc/hexanes) to give the title compound (700 mg, 67%). MS m/z: [M+H⁺]calcd for C₃₁H₃₈N₂O₃Si 515.27. found 515.5.

Preparation 246-{Benzyl-[(R)-2-(8-benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethyl]amino}hexanoicAcid{2-[(S)-3-(Carbamoyldiphenylmethyl)-pyrrolidin-1-yl]ethyl}methylamide

To a stirred solution of the product of Preparation 23 (1.57 mmol) andDIPEA (819 μL, 4.7 mmol) in acetonitrile (3.14 mL) is added the productof Preparation 22 (995 mg, 1.88 mmol). The reaction mixture is heated to80° C. for 24 h. The solvent is removed under reduced pressure and theresidue is dissolved in EtOAc (10 mL) and then washed with saturatedaqueous sodium bicarbonate solution (5 mL), dried (magnesium sulfate),and the solvent removed under reduced pressure. The crude material ispurified by column chromatography to obtain the title compound.

Preparation 256-{Benzyl-[(R)-2-(8-benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethyl]-amino}hexanoicAcid{2-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]ethyl}methylamide

To a stirred solution of the product of Preparation 25 (0.47 mmol) indichloromethane (4.7 mL) is added triethylamine trihydrofluoride (116μL, 0.71 mmol). The reaction mixture is stirred for 10 h. and then it isdiluted with dichloromethane (10 mL) and washed with saturated aqueoussodium bicarbonate solution (5 mL). The organic phase is then dried(MgSO₄) and the solvent is removed under reduced pressure to give thetitle compound.

Example 46-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]hexanoicAcid{2-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]ethyl}methylamide

To a stirred solution of the product of Preparation 25 (0.47 mmol) inethanol (4.7 mL) is added palladium (10 wt. % (dry basis) on activatedcarbon) (160 mg) and the reaction mixture is placed under an atmosphereof hydrogen and stirred overnight. The reaction mixture is then filteredand solvent is removed under reduced pressure. The crude residue ispurified by preparative HPLC to give the title compound as theditrifluoroacetate salt.

Preparation 26N-(4-Aminomethylphenyl)-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]-propionamide

To a stirred solution of 4-(N-tert-butoxycarbonylaminomethyl)aniline(756 mg, 3.4 mmol), the product of Preparation 15 (4.08 mmol) and HATU(1.55 g, 4.08 mmol) in DMF (6.8 mL) is added DIPEA (770 μL, 4.42 mmol).The reaction mixture is stirred at 50° C. overnight and then the solventis removed under reduced pressure. The resulting residue is dissolved indichloromethane (20 mL) and washed with saturated aqueous sodiumbicarbonate solution (10 mL). The organic phase is then dried (magnesiumsulfate) and the solvent is removed under reduced pressure. The crudeproduct is purified by flash chromatography. The product is dissolved inTFA/DCM (25%, 30 mL) and stirred at room temperature for 2 h. Thesolvent is then removed under reduced pressure and the crude residue isdissolved in dichloromethane (30 mL) and washed with 1N sodium hydroxide(15 mL). The organic phase is separated, dried (magnesium sulfate) andthe solvent is removed under reduced pressure to give the titlecompound.

Preparation 27N-(4-{[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]methyl}phenyl)-3-[(S)-3-(carbamoyldiphenylmethyl)-pyrrolidin-1-yl]propionamide

A solution of the product of Preparation 26 (1.04 mmol), the product ofPreparation 11 (610 mg, 1.25 mmol), sodium bicarbonate (262 mg, 3.12mmol) and sodium iodide (203 mg, 1.35 mmol) in THF (0.52 mL) are heatedat 80° C. for 12 h. The reaction mixture is diluted with dichloromethane(10 mL) and washed with saturated aqueous sodium bicarbonate solution (5mL). The organic phase is dried (MgSO₄) and the solvent is removed underreduced pressure. The crude residue is purified by flash chromatographyto give the title compound.

Preparation 28N-(4-{([(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethylamino]-methyl}phenyl)-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]propionamide

To a stirred solution of the product of Preparation 27 (0.8 mmol) indichloromethane (8 mL) is added triethylamine trihydrofluoride (261 μL,1.6 mmol). The reaction mixture is stirred for 10 h and then is dilutedwith dichloromethane (20 mL) and washed with saturated aqueous sodiumbicarbonate solution (10 mL). The organic phase is then dried (magnesiumsulfate) and the solvent is removed under reduced pressure to yield thetitle compound.

Example 53-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-N-(4-{[(R)-2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}phenyl)propionamide

To a stirred solution of the product of Preparation 28 (0.65 mmol) inethanol (6.5 mL) is added palladium (10 wt. % (dry basis) on activatedcarbon) (200 mg) and the reaction mixture is placed under a hydrogenatmosphere and stirred overnight. The reaction mixture is then filteredand the solvent is removed under reduced pressure. The crude residue ispurified by preparative HPLC to give the title compound as theditrifluoroacetate salt.

Preparation 292-[(S)-1-(2-tert-Butoxycarbonylminoethyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 1 (6.76 mmol) andDIPEA (3.54 mL, 20.3 mmol) in acetonitrile (67.6 mL) at 50° C. is added2-tert-butoxycarbonylaminoethyl bromide (1.82 g, 8.11 mmol) and thereaction mixture is heated at 50° C. overnight. The solvent is thenremoved under reduced pressure and the residue is dissolved indichloromethane (60 mL) and washed with saturated aqueous sodiumbicarbonate solution (30 mL). The organic phase is dried (magnesiumsulfate) and the solvent is removed under reduced pressure. The cruderesidue is purified by column chromatography to yield the titlecompound.

Preparation 302-[(S)-1-(2-Aminoethyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

The product of Preparation 29 is dissolved in TFA/DCM (25%, 52 mL) andstirred at room temperature for 2 h. The solvent is then removed underreduced pressure and the crude residue is dissolved in dichloromethane(30 mL) and washed with 1N sodium hydroxide (15 mL). The organic phaseis separated, dried (magnesium sulfate) and the solvent is removed underreduced pressure to give the title compound.

Preparation 314-Aminomethyl-N-{2-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]ethyl}-benzamide

To a stirred solution of the product of Preparation 30 (1 mmol),4-(tert-butoxycarbonylaminomethyl)benzoic acid (301 mg, 1.2 mmol) andHATU (456 mg, 1.2 mmol) in DMF (2 mL) is added DIPEA (226 μL, 1.3 mmol).The reaction mixture is stirred at room temperature overnight and thenthe solvent is removed under reduced pressure. The resulting residue isdissolved in dichloromethane (20 mL) and washed with saturated aqueoussodium bicarbonate solution (10 mL). The organic phase is dried(magnesium sulfate) and the solvent is removed under reduced pressure.The crude product is dissolved in TFA/DCM (25%, 10 mL) and this mixtureis stirred at room temperature for 2 h. The solvent is removed underreduced pressure and the crude residue is dissolved in dichloromethane(15 mL) and washed with 1N sodium hydroxide (5 mL). The organic phase isseparated, dried (magnesium sulfate) and the solvent is removed underreduced pressure to afford the title compound.

Preparation 324-{[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]methyl}-N-{2-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]ethyl}benzamide

A solution of the product of Preparation 32 (1.1 mmol), the product ofPreparation 11 (634 mg, 1.3 mmol), sodium bicarbonate (277 mg, 3.3 mmol)and sodium iodide (215 mg, 1.43 mmol) in THF (0.55 mL) is heated at 80°C. for 12 h. The reaction mixture is then diluted with dichloromethane(10 mL) and washed with saturated aqueous sodium bicarbonate solution (5mL). The organic phase is then dried (magnesium sulfate) and the solventis removed under reduced pressure. The crude residue is purified byflash chromatography to give the title compound.

Preparation 334-{[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethylamino]-methyl}-N-{2-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]ethyl}benzamide

To a stirred solution of the product of Preparation 32 (0.36 mmol) indichloromethane (3.6 mL) is added triethylamine trihydrofluoride (117μL, 0.72 mmol). The reaction mixture is stirred for 10 h and is thendiluted with dichloromethane (10 mL) and washed with saturated aqueoussodium bicarbonate solution (5 mL). The organic phase is dried (MgSO₄)and the solvent is removed under reduced pressure to give the titlecompound.

Example 6N-{2-[(S)-3-(Carbamoyldiphenylmethyl)pyrroidin-1-yl]ethyl}-4-{[(R)-2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}benzamide

To a stirred solution of the product of Preparation 33 (0.36 mmol) inethanol (3.6 mL) is added palladium (10 wt. % (dry basis) on activatedcarbon) (275 mg) and the reaction mixture is placed under a hydrogenatmosphere and stirred overnight. The reaction mixture is then filteredand the solvent is removed under reduced pressure. The crude residue ispurified by preparative HPLC to yield the title compound as theditrifluoroacetate salt.

Preparation 345-[(R)-2-(5-Aminopentylamino)-1-(tert-butyldimethylsilanyloxy)ethyl]-8-benzyloxy-1H-quinolin-2-one

N-tert-butoxycarbonyl-1,5-diaminopentane (1.04 g, 5.12 mmol) was addedto a solution of the product of Preparation 11 (1.00 g, 2.05 mmol) indimethyl sulfoxide (2 mL). The solution was stirred at 75° C. for 12hours, at which time LCMS analysis showed that the reaction wascomplete. The reaction mixture was then concentrated under vacuum todryness. To the residue was added dichloromethane (2 mL) andtrifluoroacetic acid (1 mL) was then added. The solution was stirred atroom temperature for about 3 hours, at which time MS analysis showedthat the reaction was complete. The solution was concentrated to halfits volume and 1N sodium hydroxide was added until the pH was adjustedto 14. The organic layer was collected, washed with brine, dried overmagnesium sulfate and then concentrated to yield 782 mg of the titlecompound as an oil. MS m/z: [M+H⁺] calcd for C₂₉H₄₃N₃O₃Si 510.8. found510.

Preparation 352-{(S)-1-[2-(3-{5-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyl-dimethylsilanyloxy)ethylamino]pentyl}ureido)ethyl]pyrrolidin-3-yl}-2,2-diphenyl-acetamide

Carbonyl diimidazole (127 mg, 0.78 mmol) is added to a solution of theproduct of Preparation 30 (78 mmol) in dimethyl formamide (4 mL) and theresulting mixture is stirred at room temperature for 3 hours. After 3hours, the product of Preparation 34 (399 mg, 0.78 mmol) is added to thereaction mixture and this mixture is stirred for 12 hours at roomtemperature. The reaction mixture is concentrated in vacuo and theresidue is diluted with ethyl acetate (5 mL). The organic layer iswashed two times with saturated sodium bicarbonate (5 mL) and then brine(5 mL). The organic layer is dried over magnesium sulfate, filtered andthen concentrated to afford the title compound.

Preparation 362-{(S)-1-[2-(3-{5-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxy-ethylamino]pentyl}ureido)ethyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

Triethylamine trihydrofluoride (0.16 mL, 1.02 mmol) is added to asolution of the product of Preparation 35 (0.68 mmol) in tetrahydrofuran(3.4 mL) and this mixture is stirred at room temperature for about 12hours. The reaction mixture is diluted with ethyl acetate (5 mL) andthis mixture is washed with 1N sodium hydroxide (5 mL), brine, driedover magnesium sulfate and concentrated to give the title compound.

Example 72-{(S)-1-[2-(3-{5-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]pentyl}ureido)ethyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

A solution of the product of Preparation 36 (0.55 mmol) in ethanol (3mL) is purged with nitrogen for about 10 minutes. Palladium (10 wt. %(dry basis) on activated carbon) (200 mg) is added and the solution isflushed again with nitrogen for about 10 minutes. The flask is purgedunder vacuum and then filled with nitrogen three times and then ahydrogen-filled balloon is placed over the flask. The reaction mixtureis stirred under hydrogen for 12 hours. The reaction mixture is thenfiltered and the organic filtrate concentrated and purified by HPLC togive the title compound as the ditrifluoroacetate salt.

Example 82-{(S)-1-[3-(3-{5-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]pentyl}ureido)propyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

Using the method described above in Preparations 34-36 and Example 7,and substituting 3-tert-Butoxycarbonylaminoprop-1-yl bromide for2-tert-butoxycarbonylaminoethyl bromide, the title compound is preparedas the ditrifluoroacetate salt.

Preparation 375-[(R)-2-[(3-Aminomethylcyclohexylmethyl)amino]-1-(tert-butyldimethylsilanyloxy)ethyl]-8-benzyloxy-1H-quinolin-2-one

A stirred solution of the product of Preparation 11 (1.46 g, 3 mmol) and1,3-cyclohexanebis(methylamine) (426 mg, 3 mmol) in DMSO (3 mL) washeated at 100° C. for 6 h. The reaction mixture was allowed to cool andit was then diluted with dichloromethane (20 mL) and washed withsaturated aqueous sodium bicarbonate solution (10 mL). The organic layerwas dried (MgSO₄) and the solvent was removed under reduced pressure.The crude residue was purified by flash chromatography (10% MeOH/DCM and0.5% NH₄OH) to give the title compound as a solid (775 mg, 50% yield).MS m/z: [M+H⁺] calcd for C₃₂H₄₇N₃O₃Si 550.3. found 550.6.

Preparation 38N-(3-{[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]methyl}cyclohexylmethyl)-3-[(S)-3-(carbamoyldiphenyl-methyl)pyrrolidin-1-yl]propionamide

To a stirred solution of the product of Preparation 37 (552 mg, 1.01mmol), the product of Preparation 15 (0.84 mmol) and HATU (384 mg, 1.01mmol) in DMF (1.68 mL) is added DIPEA (190 μL, 1.09 mmol). The reactionmixture is stirred at 50° C. overnight and then the solvent is removedunder reduced pressure. The resulting residue is dissolved indichloromethane (20 mL) and washed with saturated aqueous sodiumbicarbonate solution (10 mL). The organic phase is dried (magnesiumsulfate) and the solvent is removed under reduced pressure. The crudeproduct is purified by flash chromatography to give the title compound.

Preparation 39N-(3-{[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethylamino]-methyl}cyclohexylmethyl)-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]-propionamide

To a stirred solution of the product of Preparation 38 (0.30 mmol) indichloromethane (3 mL) is added triethylamine trihydrofluoride (98 μL,0.6 mmol). The reaction mixture is stirred for 10 h and then it isdiluted with dichloromethane (10 mL) and washed with saturated aqueoussodium bicarbonate solution (5 mL). The organic phase is dried(magnesium sulfate) and the solvent is removed under reduced pressure togive the title compound.

Example 93-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-N-(3-{[(R)-2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}cyclohexylmethyl)-propionamide

Palladium (10 wt. % (dry basis) on activated carbon) (120 mg) is addedto a stirred solution of the product of Preparation 39 (0.30 mmol) inethanol (3 mL). The reaction mixture is placed under a hydrogenatmosphere and stirred overnight. The reaction mixture is then filteredand the solvent is removed under reduced pressure. The crude residue ispurified by preparative HPLC to give the title compound as theditrifluoroacetate salt.

Preparation 40 (1R,3S)-3-Aminocyclopentanecarboxylic Acid{2-[(S)-3-(Carbamoyldiphenylmethyl)-pyrrolidin-1-yl]ethyl}amide

To a stirred solution of the product of Preparation 30 (0.94 mmol),(1R,3S)-3-tert-butoxycarbonylaminocyclopentanecarboxylic acid (258 mg,1.1 mmol) and HATU (428 mg, 1.1 mmol) in DMF (5 mL) is added DIPEA (245μL, 1.09 mmol). The reaction mixture is stirred at room temperatureovernight and then the solvent is removed under reduced pressure. Theresulting residue is dissolved in dichloromethane (20 mL) and washedwith saturated aqueous sodium bicarbonate solution (10 mL). The organiclayer is dried (magnesium sulfate) and the solvent is removed underreduced pressure. The crude product is purified by flash chromatographyand then is dissolved in a trifluoroacetic acid/DCM mixture (1 mL/5 mL)and stirred at room temperature for 1 h. The solvent is removed underreduced pressure. The residue is dissolved in dichloromethane (20 mL)and washed with 1 M sodium hydroxide (10 mL), dried (magnesium sulfate)and the solvent is removed to yield the title compound as theditrifluoroacetate salt.

Preparation 41(1R,3S)-3-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]cyclopentanecarboxylicAcid {2-[(S)-3-(Carbamoyldiphenyl-methyl)pyrrolidin-1-yl]ethyl}amide

A stirred solution of the product of Preparation 40 (0.38 mmol) and theproduct of Preparation 11 (92 mg, 0.19 mmol) in DMSO (0.38 mL) is heatedat 90° C. for 5 h. The solution is cooled and diluted with ethyl acetate(10 mL) and then washed with saturated aqueous sodium bicarbonate (5mL). The organic phase is dried (magnesium sulfate) and the solvent isremoved under reduced pressure. The crude product is purified by flashchromatography to yield the title compound.

Preparation 42(1R,3S)-3-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxy-ethylamino]cyclopentanecarboxylicAcid {2-[(S)-3-(Carbamoyldiphenylmethyl)-pyrrolidin-1-yl]ethyl}amide

To a stirred solution of the product of Preparation 41 (0.4 mmol) in THF(2 mL) is added triethylamine trihydrofluoride (130 μL, 0.8 mmol). Thereaction mixture is stirred for 10 h and is then diluted with EtOAc (10mL). The reaction mixture is washed with saturated aqueous sodiumbicarbonate solution (5 mL) and then the organic phase is dried(magnesium sulfate) and the solvent is removed under reduced pressure togive the title compound.

Example 10(1R,3S)-3-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]-cyclopentanecarboxylicAcid {2-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-ethyl}amide

To a stirred solution of the product of Preparation 42 (0.40 mmol) inethanol (3 mL) is added palladium (10 wt. % (dry basis) on activatedcarbon (120 mg). The reaction mixture is placed under a hydrogenatmosphere and stirred overnight. The reaction mixture is filtered andthe solvent is removed under reduced pressure. The crude residue ispurified by preparative HPLC to give the title compound as theditrifluoroacetate salt.

Preparation 43 4-(tert-Butoxycarbonylaminomethyl)-2-chlorophenylamine

A stirred solution of 4-aminomethyl-2-chlorophenylamine (940 mg, 6 mmol)and di-tert-butyl dicarbonate (1.44 g, 6.6 mmol) in dichloromethane (30mL) was stirred at room temperature for 4 h, at which time the reactionwas determined to be complete by LCMS. The reaction mixture was thenwashed with saturated aqueous sodium bicarbonate (15 mL) and the organiclayer was dried over sodium sulfate and the solvent was removed underreduced pressure. The resulting orange solid was recrystallized fromethyl acetate to give the title intermediate as a white solid (˜100%yield).

Preparation 44N-[4-(tert-Butoxycarbonylaminomethyl)-2-chlorophenyl]acrylamide

To a stirred solution of the product of Preparation 43 (1.54 g, 6.0mmol) in a mixture of diethyl ether (35 mL) and 1 M sodium hydroxide (35mL) was added dropwise acryloyl chloride (687 μL, 8.45 mmol). After 1 h,the organic layer was separated, dried (Na₂SO₄) and the solvent wasremoved under reduced pressure to give the title intermediate as a whitesolid (1.8 g, 96% yield).

Preparation 45N-(4-(tert-Butoxycarbonylaminomethyl)-2-chlorophenyl)-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]propionamide

A solution of the product of Preparation 1 (3.5 mmol) and the product ofPreparation 44 (1.19 g, 3.85 mmol) in a mixture of dichloromethane andmethanol (12 mL, 1:1) is heated at 60° C. for 12 h. The reaction mixtureis allowed to cool and the solvent is removed under reduced pressure.The crude material is purified by column chromatography to give thetitle intermediate.

Preparation 46N-(4-Aminomethyl-2-chlorophenyl)-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]propionamide

A solution of the product of Preparation 45 (3.3 mmol) is stirred indichloromethane (24 mL) and TFA (8 mL) for 1 h and then the solvent isremoved under reduced pressure. The crude reaction mixture is dissolvedin dichloromethane (30 mL) and washed with 1 M sodium hydroxide (2×30mL). The organic layer is dried (Na₂SO₄) and the solvent is removedunder reduced pressure to give the title intermediate.

Preparation 47N-(4-{[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]methyl}-2-chlorophenyl)-3-[(S)-3-(carbamoyldiphenyl-methyl)pyrrolidin-1-yl]propionamide

A stirred solution of the product of Preparation 46 (2.79 mmol) and theproduct of Preparation 11 (680 mg, 1.39 mmol) in DMSO (1.39 mL) isheated at 90° C. for 8 h and then cooled to room temperature. Thereaction mixture is diluted with ethyl acetate/chloroform (20 mL, 1/1)and the organic layer is washed with saturated aqueous sodiumbicarbonate (10 mL), dried (Na₂SO₄) and the solvent is removed underreduced pressure. The resulting crude residue is purified by columnchromatography to give the title intermediate.

Preparation 48N-(4-{[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethylamino]-methyl}-2-chlorophenyl)-3-[(S)-3-(carbamoyldiphenylmethyl)pyrrolidin-1-yl]-propionamide

To a stirred solution of the product of Preparation 47 (1.23 mmol) indichloromethane (12 mL) is added Et₃N.3HF (401 μL, 0.6 mmol). Thereaction mixture is stirred for 10 h and then diluted withdichloromethane (10 mL). This mixture is washed with saturated aqueoussodium bicarbonate solution (5 mL) and the organic layer dried (Na₂SO₄)and the solvent is removed under reduced pressure to give the titleintermediate.

Example 113-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-N-(2-chloro-4-{[(R)-2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}phenyl)-propionamide

To a stirred solution of the product of Preparation 48 (1.2 mmol) inethanol (12 mL) is added Pd/C (290 mg) and the reaction mixture isplaced under a hydrogen atmosphere and stirred overnight. The reactionmixture is then filtered and the solvent is removed under reducedpressure. The crude residue is purified by preparative HPLC to give thetitle compound as the ditrifluoroacetate salt.

Preparation 49 2-Chloroethanesulfonic Acid(5-tert-Butoxycarbonylaminopentyl)amide

To a stirred solution of 5-(tert-butoxycarbonylamino)pentylamine (1.00g, 4.94 mmol) and triethylamine (689 μL g, 4.94 mmol) in dichloromethane(22 mL) at 0° C. was added 2-chloro-1-ethanesulfonyl chloride (470 μL,4.50 mmol). The reaction mixture was stirred for 2 h at room temperatureand then washed with saturated aqueous sodium bicarbonate solution (15mL). The organic layer was dried (Na₂SO₄) and the solvent was removedunder reduced pressure to give the title compound (100% yield), whichwas used in the next step without further purification.

Preparation 502-{(S)-1-[2-(5-tert-Butoxycarbonylaminopentylsulfamoyl)ethyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

A solution of the product of Preparation 1 (3.5 mmol) and the product ofPreparation 49 (1.62 g, 4.94 mmol) in dichloromethane and methanol (22mL, 1:1) is heated at 60° C. for 5 h. The reaction mixture is allowed tocool to room temperature and the solvent is removed under reducedpressure. The crude residue is dissolved in dichloromethane (20 mL) andwashed with saturated aqueous sodium bicarbonate solution (10 mL). Theorganic layer is then dried (Na₂SO₄) and solvent is removed underreduced pressure. The crude residue is purified by column chromatographyto give the title intermediate.

Preparation 512-{(S)-1-[2-(5-Aminopentylsulfamoyl)ethyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

A solution of the product of Preparation 50 (2.72 mmol) is stirred indichloromethane (21 mL) and TFA (7 mL) for 1 h and then the solvent isremoved under reduced pressure. The crude reaction mixture is dissolvedin dichloromethane (30 mL) and washed with 1 M sodium hydroxide (2×30mL). The organic layer is dried (Na₂SO₄) and the solvent is then removedunder reduced pressure to give the title intermediate.

Preparation 522-[(S)-1-(2-{5-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyl-dimethylsilanyloxy)ethylamino]pentylsulfamoyl}ethyl)pyrrolidin-3-yl]-2,2-diphenyl-acetamide

A stirred solution of the product of Preparation 51 (1.88 mmol) and theproduct of Preparation 11 (460 mg, 0.94 mmol) in DMSO (0.92 mL) isheated at 90° C. for 8 h and then cooled to room temperature. Thereaction mixture is diluted with ethyl acetate/chloroform (20 mL, 1/1)and the organic layer is washed with saturated aqueous sodiumbicarbonate solution (10 mL), dried (Na₂SO₄) and the solvent is removedunder reduced pressure. The resulting crude residue is purified bycolumn chromatography to give the title intermediate.

Preparation 532-[(S)-1-(2-{5-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxy-ethylamino]pentylsulfamoyl}ethyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 52 (0.56 mmol) indichloromethane (5.6 mL) is added triethylamine trihydrofluoride (183μL, 1.12 mmol). The reaction mixture is stirred for 10 h anddichloromethane (10 mL) is added. The resulting mixture is washed withsaturated aqueous sodium bicarbonate solution (5 mL). The organic layeris dried (Na₂SO₄) and the solvent is removed under reduced pressure togive the title intermediate.

Example 122-[(S)-1-(2-{5-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]pentylsulfamoyl}ethyl)pyrrolidin-3-yl]-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 53 (0.56 mmol) inethanol/methanol (5.6 mL, 1/1) is added Pd/C (131 mg) and the reactionmixture is placed under a hydrogen atmosphere and stirred overnight. Thereaction mixture is then filtered and the solvent is removed underreduced pressure. The crude residue is purified by preparative HPLC togive the title compound as the ditrifluoroacetate salt.

Preparation 542-((S)-{2-[(4-Formylbenzenesulfonyl)methylamino]ethyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 21 (1 mmol) andtriethyl amine (167 μL, 1.2 mmol) in dichloromethane (5 mL) is added4-formylbenzenesulfonyl chloride (225 mg, 1.1 mmol). After 1 h at roomtemperature, the reaction mixture is washed with saturated aqueoussodium bicarbonate solution (5 mL). The organic layer is then dried(Na₂SO₄) and solvent is removed under reduced pressure to give the titleintermediate.

Preparation 552-((S)-1-{2-[(4-{[(R)-2-(tert-Butyldimethylsilanyloxy)-2-(8-hydroxy-2-oxo-1,2-dihydro-quinolin-5-yl)ethylamino]methyl}benzenesulfonyl)methylamino]ethyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

A solution of5-[(R)-2-amino-1-(tert-butyldimethylsilanyloxy)ethyl]-8-hydroxy-1H-quinolin-2-one (293 mg, 0.74 mmol) and the product of Preparation 54in dichloromethane and methanol (6.2 mL, 1/1) is stirred at roomtemperature for 1 h and then sodium triacetoxyborohydride (394 mg, 1.86mmol) is added. The reaction mixture is stirred for 4 h and thenacidified with concentrated hydrochloric acid and the solvent is removedunder reduced pressure to provide the title compound.

Example 132-((S)-1-{2-[(4-{[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]methyl}benzenesulfonyl)methylamino]ethyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

A stirred solution of the product of Preparation 55 (0.62 mmol) in 1Mhydrochloric acid (5 mL) and acetonitrile (5 mL) is heated at 60° C. for8 h. The reaction mixture is cooled to room temperature and the solventis removed under reduced pressure. The crude residue is purified bypreparative HPLC to give the title compound as the ditrifluoroacetatesalt.

Preparation 56 (3-Aminomethylphenyl)methanol Hydrochloride

(a) (3-tert-Butoxycarbonylmethylphenyl)methanol

Borane dimethyl sulfide (2.05 mL, 21.6 mmol) was added to a solution of3-(tert-butoxycarbonylaminomethyl)benzoic acid (1.81 g, 7.20 mmol) intetrahydrofuran (24 mL). and the resulting mixture was stirred at roomtemperature for 3 hours. The reaction mixture was then diluted withethyl acetate (20 mL) and the layers were separated. The organic layerwas washed with saturated sodium bicarbonate, saturated sodium chloride,dried over magnesium sulfate and concentrated to give the title compoundas a yellow oil (1.71 g).

(b) (3-Aminomethylphenyl)methanol Hydrochloride

To the product of step (a) (1.71 g, 7.2 mmol) was added a solution of 4M hydrochloric acid in dioxane (9 mL, 36 mmol) and the resulting mixturewas stirred at room temperature for 1 h. The reaction mixture was thenconcentrated and the residue was diluted with diethyl ether (50 mL) andfiltered to provide the title compound as a white solid (1.09 g).

Preparation 572-((S)-1-{2-[3-(3-Hydroxymethylbenzyl)ureido]ethyl}pyrrolidin-3-yl)-2,2-diphenyl-acetamide

A 0.2 M solution of the product of Preparation 30 (2.24 mmol) inN,N-dimethylformamide is added dropwise to a solution of1,1-carbonyldiimidazole (364 mg, 2.24 mmol) and diisopropylethylamine(0.31 mL, 2.24 mmol) in N,N-dimethylformamide (11 mL) and the resultingmixture is stirred at room temperature for 2 h. Diisopropylethylamine(0.31 mL, 2.24 mmol) and the product of Preparation 56 (578 mg, 3.4mmol) are added and this mixture is stirred at 50° C. for 12 hours. Thereaction mixture is then concentrated to dryness and the residue isdiluted with dichloromethane (20 mL) and this solution is washed withsaturated sodium bicarbonate (2×), saturated sodium chloride, dried overmagnesium sulfate, and concentrated to provide the title compound.

Preparation 582-((S)-1-{2-[3-(3-Formylbenzyl)ureido]ethyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

A solution of the product of Preparation 57 (2.23 mmol) indichloromethane (11.1 mL) is cooled to 0° C. and diisopropylethylamine(1.17 mL, 6.70 mmol) and dimethyl sulfoxide (0.949 mL, 13.4 mmol) areadded. After about 10 minutes, pyridine sulfur trioxide complex (1.06 g,6.70 mmol) is added and the resulting mixture is stirred at 0° C. for 2h. The reaction is then quenched with water (15 mL) and the organiclayer is washed with cold water (3×), dried over magnesium sulfate andconcentrated to provide the title compound.

Preparation 592-((S)-1-{2-[3-(3-{[(R)-2-(tert-Butyldimethylsilanyloxy)-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}benzyl)ureido]ethyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

5-[(R)-2-Amino-1-(tert-butyldimethylsilanyloxy)ethyl]-8-hydroxy-1H-quinolin-2-one(575 mg, 1.40 mmol) is added to a solution of the product of Preparation58 (1.2 mmol) and diisopropylamine (0.25 mL, 1.40 mmol) indichloromethane (6 mL) and the resulting mixture is stirred at roomtemperature for 45 min. Sodium triactetoxyborohydride (385 mg, 1.80mmol) is then added and this mixture is stirred at room temperature for12 h. The reaction is then quenched with 10% aqueous hydrochloric acid(5 mL) and the layers are separated. The organic layer is washed withsaturated sodium bicarbonate, saturated sodium chloride, dried overmagnesium sulfate and concentrated to give the title compound.

Example 142-((S)-1-{2-[3-(3-{[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]methyl}benzyl)ureido]ethyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

Triethylamine trihydrofluoride (2.4 mL, 13.6 mmol) is added to asolution of the product of Preparation 59 (1.36 mmol) in dichloromethane(2 mL) and the resulting mixture is stirred at room temperature for 15h. The reaction mixture is then concentrated under vacuum to dryness andthe residue is dissolved in a 1:1 mixture of water and acetonitrile with0.1% TFA and this mixture is purified by HPLC to provide the titlecompound as the ditrifluoroacetate salt.

Preparation 602-{(S)-1-[(E)-3-(4-Nitrophenyl)allyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

The product of Preparation 1 (0.01 mol) and p-nitrocinnamaldehyde (1.77g, 0.01 mol) are stirred in 50 mL of dichloromethane for 2 h. Sodiumtriacetoxyborohydride (6.33 g, 0.03 mol) is added and the resultingmixture is stirred for 2 h. The reaction is then quenched with 10 mL ofwater and this mixture is diluted with dichloromethane (100 mL). Theorganic layer is washed with saturated sodium bicarbonate (2×), brine,dried over Na₂SO₄, filtered and concentrated to provide the titlecompound.

Preparation 612-{(S)-1-[3-(4-Aminophenyl)propyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

The product of Preparation 60 (5.4 mmol) is dissolved in 100 mL ofethanol and the resulting solution is purged with nitrogen for 30 min.Palladium on carbon (2.5 g; 50% w/w water; 10% Pd; 1.1 mmol Pd) is thenadded while degassing with nitrogen. This mixture is then placed underhydrogen (50 psi) until hydrogen is no longer consumed (˜30 minutes).The mixture is then purged with nitrogen, filtered through Celite andconcentrated. The residue is dissolved in ethyl acetate and this mixtureis washed with saturated sodium bicarbonate (2×), brine, dried (Na₂SO₄),filtered and concentrated to provide the title compound.

Preparation 622-((S)-1-{3-[4-(4-{2-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-(tert-butyldimethylsilanyloxy)ethylamino]ethyl}phenylamino)phenyl]propyl}-pyrrolidin-3-yl)-2,2-diphenylacetamide

To a 25 mL round-bottomed flask is added the product of Preparation 61(0.8 mmol);8-benzyloxy-5-[(R)-2-[2-(4-bromophenyl)ethylamino]-1-(tert-butyldimethylsilanyloxy)ethyl]-1H-quinolin-2-one(769 mg, 1.2 mmol); tris(dibenzylideneacetone)dipalladium(0) (73 mg,0.08 mmol, 20% Pd); and 2-(dicyclohexylphosphino)biphenyl (84 mg, 0.24mmol). This mixture is purged with nitrogen and then dry, degassedtoluene (8 mL, 0.1 M) is added and the resulting mixture is heated at70° C. for 30 min. Sodium tert-butoxide (382 mg, 4.0 mmol) is thenadded, and the temperature is raised to 95° C. for 4 h. The reactionmixture is then cooled to room temperature and diluted with ethylacetate. This mixture is washed with saturated sodium bicarbonate (2×),brine, dried (Na₂SO₄), filtered and concentrated to provide the titlecompound.

Preparation 632-((S)-1-{3-[4-(4-{2-[(R)-2-(8-Benzyloxy-2-oxo-1,2-dihydroquinolin-5-yl)-2-hydroxyethylamino]ethyl}phenylamino)phenyl]propyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

The product of Preparation 62 is dissolved in dichloromethane (10 mL)and triethylamine trihydrofluoride (10 eq.) is added. The reactionmixture is stirred overnight and then diluted with dichloromethane andthe organic layer is washed with saturated sodium bicarbonate (2×),brine, dried (Na₂SO₄), filtered and concentrated to provide crudeproduct. This material is purified by silica gel chromatography toprovide the title compound.

Example 152-((S)-1-{3-[4-(4-{2-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]ethyl}phenylamino)phenyl]propyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

The product of Preparation 63 is dissolved in 10 mL of ethanol and thismixture is purged with nitrogen for 15 minutes. Palladium on carbon (10%Pd, 50% w/w water, 0.2 eq. Pd) is added while degassing. The resultingmixture is then placed under 1 atm. of hydrogen for 2 h. The solution isthen purged with nitrogen for 15 min and then filtered through Celiteand concentrated. The resulting residue is purified by prep HPLC toafford the title compound as the ditrifluoroacetate salt.

Preparation 642-{(S)-1-[2-Fluoro-3-(4-hydroxymethylpiperidin-1-ylmethyl)benzyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

The product of Preparation 1 (1.69 mmol),2,6-bis(bromomethyl)-1-fluorobenzene (476 mg, 1.69 mmol,piperidin-4-ylmethanol (195 mg, 1.69 mmol) and potassium carbonate (466mg, 3.37 mmol) are suspended in acetonitrile (5 mL) and stirred at roomtemperature for 18 h. The reaction mixture is then concentrated and theresidue is dissolved in dichloromethane/water. The layers are separatedand the organic layer is washed with water (2×), brine, dried (MgSO₄)and concentrated. The crude material is purified by silica gel columnchromatography to give the title compound.

Preparation 652-{(S)-1-[2-Fluoro-3-(4-formylpiperidin-1-ylmethyl)benzyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

The product of Preparation 64 (0.53 mmol) is dissolved indichloromethane and to this mixture is added diisopropylethylamine (280μL, 1.6 mmol) and dimethyl sulfoxide (115 μL, 1.6 mmol). The reactionmixture is cooled to −15° C. under nitrogen and pyridine sulfur trioxidecomplex (255 mg, 1.6 mmol) is added and the resulting mixture is stirredfor 40 min. The reaction is then quenched with water and the layers areseparated. The organic layer is washed with aqueous NaH₂PO₄ (1 M×3),brine, dried (MgSO₄) and concentrated to provide the title compound.

Example 162-{(S)-1-[2-Fluoro-3-(4-{[(R)-2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}piperidin-1-ylmethyl)benzyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

The product of Preparation 65 (0.48 mmol) is dissolved in a 1:1 mixtureof dichloromethane and methanol (6 mL) and to this mixture is added5-[(R)-2-amino-1-(tert-butyldimethylsilanyloxy)ethyl]-8-hydroxy-1H-quinolin-2-oneacetate (228 mg, 0.58 mmol) and sodium triacetoxyborohydride (317 mg,1.5 mmol). The reaction mixture is stirred under nitrogen at roomtemperature for 18 h and then concentrated. The residue is dissolved ina 2:3 mixture of acetonitrile and aqueous 6 N hydrochloric acid, andthis mixture is heated at 55° C. for 4 hours. The reaction mixture isthen concentrated and the residue is dissolved inwater/acetonitrile/trifluoroacetic acid (1:1:0.005) and purified byreverse phase column chromatography to afford the title compound as theditrifluoroacetate salt.

Preparation 66 2-[4-(3-Bromopropoxy)phenyl]ethanol

To a solution of 4-hydroxyphenethyl alcohol (4.37 g, 31.0 mmol) andpotassium carbonate (6.55 g, 47.0 mmol) in acetonitrile (62.0 mL) wasadded 1,3 dibromopropane (31.0 mL, 316 mmol). The reaction mixture washeated to 70° C. for 12 hours and then cooled to room temperature,filtered and concentrated under vacuum. The resulting oil was purifiedby silica gel chromatography using a mixture of 4:1 hexanes and ethylacetate to give the title compound (6.21 g) as a white solid.

Preparation 672-((S)-1-{3-[4-(2-Hydroxyethyl)phenoxy]propyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

To a solution of the product of Preparation 66 (1.11 g, 4.30 mmol) anddiisopropylethylamine (0.90 mL, 5.10 mmol) in acetonitrile (21.5 mL) isadded the product of Preparation 1 (4.30 mmol) and the resulting mixtureis stirred at 60° C. for 12 h. The reaction mixture is then diluted withdichloromethane (20 mL) and washed with saturated sodium bicarbonate (25mL), saturated sodium chloride (25 mL), dried over magnesium sulfate andconcentrated to provide the title compound.

Preparation 682-((S)-1-{3-[4-(2-Oxoethyl)phenoxy]propyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

A solution of the product of Preparation 67 (1.53 mmol) anddichloromethane (75 mL) is cooled to about 5° C. anddiisopropylethylamine (798 mL, 4.58 mmol) and dimethyl sulfoxide (649mL, 9.15 mmol) are added. Pyridine sulfur trioxide (728 mg, 4.58 mmol)is then added and the resulting mixture is stirred at 5° C. for 45 min.The reaction mixture is then diluted with dichloromethane (20 mL) andwashed with saturated sodium bicarbonate (25 mL), saturated sodiumchloride (25 mL), dried over magnesium sulfate and concentrated toprovide the title compound.

Preparation 692-{(S)-1-[3-(4-{2-[(R)-2-(tert-Butyldimethylsilanyloxy)-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]ethyl}phenoxy)propyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

The product of Preparation 68 (1.28 mmol) is dissolved in methanol (6.4mL) and5-[(R)-2-amino-1-(tert-butyldimethylsilanyloxy)ethyl]-8-hydroxy-1H-quinolin-2-one(605 mg, 1.53 mmol) and diisopropylethylamine (0.27 mL, 1.53 mmol) areadded. Sodium triacetoxyborohydride (405 mg, 1.91 mmol) is then addedand the reaction mixture is stirred at room temperature for 3 h. Thereaction mixture is then concentrated to dryness and the residue isdiluted with ethyl acetate (20 mL) and this solution is washed withsaturated sodium bicarbonate (25 mL), saturated sodium chloride (25 mL),dried over magnesium sulfate and concentrated to give the titlecompound.

Example 172-{(S)-1-[3-(4-{2-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]ethyl}phenoxy)propyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

Triethylamine trihydrofluoride (1.5 mL, 8.87 mmol) is added to asolution of the product of Preparation 69 (0.89 mmol) in dichloromethane(4.5 mL) and the resulting mixture is stirred at room temperature for 24h. The mixture is then concentrated under vacuum and purified by HPLC togive the title compound as the ditrifluoroacetate salt.

Preparation 70 Methyl 4-Iodophenylacetate

To a stirred solution of 4-iodophenylacetic acid (5.0 g, 19.1 mmol) inMeOH (200 mL) was added 4N hydrochloric acid in dioxane (10 mL). Thereaction mixture was stirred for 24 h at room temperature and then thesolvent was removed under reduced pressure to give the title compound(5.17 g, 98% yield), which was used without further purification.

Preparation 71 Methyl [4-(4-Hydroxybut-1-ynyl)phenyl]acetate

To a stirred solution of the product of Preparation 70 (4.5 g, 16.3mmol) in diethylamine (100 mL) was added but-3-yn-1-ol (1.9 mL, 32.6mmol), Pd(PPh₃)₂Cl₂ (500 mg, 1.63 mmol) and CuI (154 mg, 0.815 mmol) andresulting mixture was stirred for 17 h at room temperature. The solventwas then removed under reduced pressure and the residue was dissolved indiethyl ether (200 mL) and this solution was filtered to remove salts.The solvent was then removed under reduced pressure and the crudeproduct was purified by silica gel chromatography (60% EtOAc/Hexane) toafford the title intermediate (3.03 g, 91% yield).

Preparation 72 Methyl [4-(4-Hydroxybutyl)phenyl]acetate

A stirred solution of the product of Preparation 71 (2.8 g, 12.8 mmol)in methanol (50 mL) was flushed with nitrogen and then 10% palladium oncarbon (400 mg, 20% wt/wt) was added. The reaction flask was thenalternately placed under vacuum and flushed with hydrogen for cycles andthen stirred under hydrogen for 14 h. The reaction mixture was flushedwith nitrogen and then filtered and the solvent removed under reducedpressure to give the title compound (2.75 g, 97% yield), which was usedwithout further purification.

Preparation 73 Methyl(4-{4-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]butyl}-phenyl)acetate

(a) Methyl {4-[4-(Toluene-4-sulfonyloxy)butyl]phenyl}acetate

To a stirred solution of the product of Preparation 72 (2.6 g, 12.5mmol) in THF (100 mL) was added DABCO (2.6 g, 25.0 mmol) and thenp-toluenesulfonyl chloride (2.44 g, 13.75 mmol). The reaction mixturewas stirred at room temperature for 23 h and then solvent was removedunder reduced pressure and the residue was dissolved in dichloromethane(200 mL). The organic layer was then washed with water (2×100 mL), 1Nhydrochloric acid (100 mL), aqueous saturated sodium chloride solution(100 mL), dried (MgSO₄), filtered and the solvent removed under reducedpressure to give the title compound, which was used without furtherpurification.

(b) Methyl(4-{4-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]butyl}-phenyl)acetate

To the crude product from step (a) is added DMF (50 mL),diisopropylethylamine (3.0 mL, 17.3 mmol) and the product of Preparation1 (8.1 mmol). The reaction mixture is stirred at room temperature for 18h and then the solvent is removed under reduced pressure to give thetitle compound.

Preparation 742-((S)-1-{4-[4-(2-Hydroxyethyl)phenyl]butyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 73 (4.0 mmol) in THF(100 mL) is added dropwise DIBAL (24 mL, 24 mmol, 1.0 M in THF). Afterthe addition is complete, the reaction mixture is stirred for 3 h andthen quenched by slow addition of methanol (until gas evolution ceased).The mixture is then stirred for 30 min. and then ethyl acetate (200 mL)and aqueous 1N sodium hydroxide (200 mL) are added. The organic layer isseparated and washed with aqueous saturated sodium chloride solution(100 mL), dried (MgSO₄), filtered and the solvent is removed underreduced pressure to give the title compound.

Example 182-{(S)-1-[4-(4-{2-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]ethyl}phenyl)butyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 74 (1.06 mmol) indichloromethane (25 mL) is added dimethyl sulfoxide (0.60 mL, 10.6 mmol)and diisopropylethylamine (0.921 mL, 5.3 mmol). The reaction mixture isthen cooled to −10° C. and pyridine sulfur trioxide (842 mg, 5.3 mmol)is added. The reaction mixture is stirred for 1 h and then quenched byadding water (100 mL). This mixture is stirred for 10 min and then theorganic layer is removed and washed with aqueous saturated sodiumchloride solution (100 mL), dried (MgSO₄) and then filtered. To thefiltrate is added methanol (25 mL),5-[(R)-2-amino-1-(tert-butyldimethylsilanyloxy)ethyl]-8-hydroxy-1H-quinolin-2-oneacetate (419 mg, 1.06 mmol) and sodium triacetoxyborohydride (468 mg,2.12 mmol). This mixture is stirred for 16 h and then condensed and, tothe resulting mixture, is added a 1:1 mixture of acetonitrile andaqueous 4N hydrochloric acid (20 mL). This mixture is heated at 50° C.for 17 h and then the solvent is removed under reduced pressure. To theresidue is added a 1:1 mixture of acetic acid and water (8.0 mL) and themixture is chromatographed on reverse-phase silica gel to afford thetitle compound as the ditrifluoroacetate salt.

Preparation 75 Ethyl 3-[5-(2-Ethoxycarbonylvinyl)thiophen-2-yl]acrylate

To a stirred solution of sodium hydride (2.1 g, 53 mmol, 60% in mineraloil) in THF (200 mL) was slowly added triethylphosphonoacetate (10 mL,50 mmol) Hydrogen gas evolution was observed and the reaction wasstirred until gas evolution ceased (about 30 min). To this reactionmixture was added 2,5-thiophenedicarboxaldehyde (3 g, 21 mmol) and thereaction mixture was stirred for 1 h. The solvent was removed underreduced pressure and the residue was dissolved in dichloromethane (200mL). The organic layer was washed with water (100 mL), aqueous 1Nhydrochloric acid (100 mL), aqueous saturated sodium chloride solution(100 mL), dried (MgSO₄), filtered and the solvent removed under reducedpressure to give the title compound (5.8 g, 98% yield), which was usedwithout further purification.

Preparation 76 Ethyl3-[5-(2-Ethoxycarbonylethyl)thiophen-2-yl]propionate

A stirred solution of the product of Preparation 75 (5.8 g, 21 mmol) inmethanol (200 mL) was flushed with nitrogen and 10% palladium on carbon(576 mg, 10% wt/wt) was added. The reaction flask was alternately placedunder vacuum and flushed with hydrogen for 3 cycles and then thereaction mixture was stirred under hydrogen for 1 h. The mixture wasthen was flushed with nitrogen, filtered and the solvent removed underreduced pressure to give the title compound (5.8 g, 99% yield), whichwas used without further purification.

Preparation 77 3-[5-(3-Hydroxypropyl)thiophen-2-yl]propan-1-ol

To a stirred solution of DIBAL (88 mL, 88 mmol, 11.0M in cyclohexane) inTHF (300 mL) at −78° C. was added dropwise the product of Preparation 76(5.0 g, 17.6 mmol). After the addition was complete, the reactionmixture was warmed to room temperature over 30 min and then quenched byslow addition of aqueous 1N hydrochloric acid (200 mL). Dichloromethane(400 mL) was added and the layers were separated. The aqueous layer waswashed with dichloromethane (4×100 mL) and the combined organic layerswere washed with aqueous saturated sodium chloride solution (100 mL),dried (MgSO₄), filtered and the solvent removed under reduced pressureto give the title compound (3.0 g, 85% yield), which was used withoutfurther purification.

Preparation 782-((S)-1-{3-[5-(3-Hydroxypropyl)thiophen-2-yl]propyl}pyrrolidin-3-yl)-2,2-diphenyl-acetamide

(a) Toluene-4-sulfonic Acid 3-[5-(3-Hydroxypropyl)thiophen-2-yl]propylEster

To a stirred solution of the product of Preparation 90 (423 mg, 2.1mmol) in THF (20 mL) was added DABCO (420 mg, 4.2 mmol) and thenp-toluenesulfonyl chloride (442 mg, 2.3 mmol). The reaction mixture wasstirred at room temperature for 2 h and then the solvent was removedunder reduced pressure and the residue was dissolved in dichloromethane(200 mL). The organic layer was washed with water (2×100 mL), aqueoussaturated sodium chloride solution (100 mL), dried MgSO₄), filtered andthe solvent removed under reduced pressure to give the title compound,which was used without further purification.

(b)2-((S)-1-{3-[5-(3-Hydroxypropyl)thiophen-2-yl]propyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

To the product from step (a) is added acetonitrile (20 mL),diisopropylethylamine (0.5 mL, 2.8 mmol) and the product of Preparation1 (2.11 mmol). The reaction mixture is heated to 50° C. for 20 h andthen cooled to room temperature and the solvent is removed under reducedpressure. The residue is purified by silica gel chromatography to affordthe title compound.

Preparation 792-{(S)-1-[3-(5-{3-[(R)-2-(tert-Butyldimethylsilanyloxy)-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]propyl}thiophen-2-yl)propyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 78 (0.94 mmol) indichloromethane (20 mL) is added dimethyl sulfoxide (0.21 mL, 3.7 mmol)and diisopropylethylamine (0.65 mL, 3.7 mmol). This mixture is cooled to−10° C. and pyridine sulfur trioxide (444 mg, 2.8 mmol) is added. Thereaction mixture is stirred for 3 h and then is quenched by adding water(100 mL). This mixture is stirred for 10 min and then the organic layeris removed and is washed with aqueous saturated sodium chloride solution(100 mL), dried (MgSO₄) and filtered. To the filtrate is added methanol(20 mL),5-[(R)-2-amino-1-(tert-butyldimethylsilanyloxy)ethyl]-8-hydroxy-1H-quinolin-2-oneacetate (368 mg, 0.93 mmol) and then sodium triacetoxyborohydride (412mg, 1.86 mmol). This mixture is stirred for 19 h and then the mixture iscondensed to give the title compound.

Example 192-{(S)-1-[3-(5-{3-[(R)-2-Hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)-ethylamino]propyl}thiophen-2-yl)propyl]pyrrolidin-3-yl}-2,2-diphenylacetamide

To the crude product from Preparation 79 is added a 1:1 mixture ofacetonitrile and aqueous 4 N hydrochloric acid (25 mL). This mixture isheated at 50° C. for 17 h and then the solvent is then removed underreduced pressure. To the residue is added a 1:1 mixture of acetic acidand water (8.0 mL) and this mixture is chromatographed on reverse-phasesilica gel to afford the title compound as the ditrifluoroacetate salt.

Preparation 80 Methyl 4-Amino-5-chloro-2-methoxybenzoate

To a solution of 4-amino-5-chloro-3-methoxybenzoic acid (1.008 g, 5.0mmol) in a mixture of toluene (9 mL) and methanol (1 mL) at 0° C. wasadded (trimethylsilyl)diazomethane (2.0 M in hexane, 3.0 mL, 6.0 mmol)dropwise. The reaction mixture was then warmed to room temperature andstirred for 16 h. Excess (trimethylsilyl)diazomethane was quenched byadding acetic acid until the bright yellow color of the reaction mixturedisappeared. The mixture was then concentrated in vacuo to give thetitle compound as an off-white solid, which was used without furtherpurification.

Preparation 81 Methyl 4-Acryloylamino-5-chloro-2-methoxybenzoate

To crude product of Preparation 93 was added dichloromethane (10 mL, 0.5M) and triethylamine (2.1 mL, 15 mmol). This mixture was cooled to 0° C.and acryloyl chloride (812 μL, 10 mmol) was added dropwise withstirring. After 2 h, the reaction was quenched by adding methanol (about2 mL) at 0° C. and the resulting mixture was stirred at room temperaturefor 15 min and then concentrated in vacuo. Dichloromethane (30 mL) andwater (30 mL) were added to the residue and this mixture was mixedthoroughly. The layers were separated and the aqueous layer wasextracted with dichloromethane (20 mL). The organic layers werecombined, dried (Na₂SO₄), filtered and the solvent was removed in vacuoto give the title compound as a brown foamy solid.

Preparation 82 Methyl4-{3-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]propionylamino}-5-chloro-2-methoxybenzoate

To the crude product from Preparation 81 is added the product ofPreparation 1 (4.5 mmol) and a mixture of THF (22.5 mL) and methanol(2.5 mL). This mixture is heated at 50° C. with stirring for 16 h andthen the solvent is removed in vacuo. The residue is chromatographed togive the title compound.

Preparation 833-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-N-(2-chloro-4-hydroxymethyl-5-methoxyphenyl)propionamide

To a solution of the product of Preparation 82 (1.45 mmol) in a mixtureof THF (4.5 mL) and methanol (0.5 mL) at 0° C. is added lithiumborohydride (32 mg, 1.45 mmol). The reaction mixture is allowed to warmto room temperature and is stirred for 41 h. The reaction is thenquenched by adding 1 N aqueous hydrochloric acid at 0° C. until no morebubbling is observed and this mixture is stirred for 10 min. The solventis removed in vacuo and the residue is dissolved in acetonitrile (about2 mL). This solution is purified by prep-RP-HPLC. The appropriatefractions are collected and combined and lyophilized to give the titlecompound as a trifluoroacetate salt. This salt is treated with isopropylacetate (10 mL) and 1N aqueous sodium hydroxide (10 mL) and the organiclayer is collected, dried (Na₂SO₄), filtered and the solvent is removedin vacuo to give the title compound.

Preparation 843-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-N-(2-chloro-4-formyl-5-methoxyphenyl)propionamide

To a solution of the product of Preparation 83 (0.3 mmol) indichloromethane (3 mL) is added dimethyl sulfoxide (213 μL, 3.0 mmol)and diisopropylethylamine (261 μL, 1.5 mmol). This mixture is cooled to−20° C. and sulfur trioxide pyridine complex (238 mg, 1.5 mmol) is addedslowly. After 30 min, the reaction mixture is quenched by adding water(about 3 mL). The layers are separated and the organic layer is dried(Na₂SO₄), filtered and the solvent is removed in vacuo to give the titlecompound.

Preparation 85N-(4-{[(R)-2-(tert-Butyldimethylsilanyloxy)-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}-2-chloro-5-methoxyphenyl)-3-[(S)-3-(carbamoyldiphenyl-methyl)pyrrolidin-1-yl]propionamide

To the product from Preparation 84 in a mixture of dichloromethane (0.5mL) and methanol (0.5 mL) is added5-[(R)-2-amino-1-(tert-butyldimethylsilanyloxy)ethyl]-8-hydroxy-1H-quinolin-2-oneacetate (124.1 mg, 3.1 mmol) and the resulting mixture is stirred atroom temperature for 1.5 h. Sodium triacetoxyborohydride (190.7 mg, 0.9mmol) is added and the resulting mixture is stirred at room temperaturefor 15 h. The reaction is quenched by adding water (about 0.2 mL) andthe mixture is concentrated in vacuo to give the title compound.

Example 203-[(S)-3-(Carbamoyldiphenylmethyl)pyrrolidin-1-yl]-N-(2-chloro-4-{[(R)-2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl)ethylamino]methyl}-5-methoxyphenyl)-propionamide

To a suspension of the product of Preparation 85 in dichloromethane (1.0mL, 0.3 M) is added triethylamine trihydrofluoride (245 μL, 1.5 mmol).This mixture is stirred at room temperature for 45 h and then themixture is concentrated in vacuo. The residue is dissolved in a mixtureof DMF (0.5 mL), acetonitrile/water (1:1, with 0.1% TFA, 0.6 mL), TFA(0.3 mL) and acetonitrile (about 1 mL) and this mixture is purified byprep-RP-HPLC. The appropriate fractions are collected and combined andlyophilized to give the title compound as the ditrifluoroacetate salt.

Examples 21-150

Using the methods described above and the appropriate startingmaterials, the compounds of Examples 21 to 150 as shown in Table I areprepared.

Preparation 86N-{2-Benzyloxy-5-[(R)-2-bromo-1-(tert-butyldimethylsilanyloxy)ethyl]phenyl}-formamide

(R)-2-Bromo-1-(3-formamido-4-benzyloxyphenyl)ethanol (9.9 g, 28 mmol)was dissolved in dimethylformamide (36 mL). Imidazole (2.3 g, 34 mmol)and tert-butyldimethylsilyl chloride (4.7 g, 31 mmol) were added. Thesolution was stirred under nitrogen atmosphere for 72 h. Additionalimidazole (0.39 g, 5.7 mmol) and tert-butyldimethylsilyl chloride (0.64g, 4.3 mmol) were added and the reaction was stirred for an additional20 h. The reaction mixture was then diluted with a mixture of isopropylacetate (53 mL) and hexanes (27 mL) and transferred to a separatoryfunnel. The organic layer was washed twice with a mixture of water (27mL) and saturated aqueous sodium chloride (27 mL) followed by a finalwash with saturated aqueous sodium chloride (27 mL). The organic layerwas dried over sodium sulfate. Silica gel (23.6 g) and hexanes (27 mL)were added and the suspension was stirred for 10 min. The solids wereremoved by filtration and the filtrate concentrated under vacuum. Theresidue was crystallized from hexanes (45 mL) to afford 8.85 g (19 mmol,68%) of the title compound as a solid. MS m/z: [M+H⁺] calcd forC₂₂H₃₀NO₃SiBr 464.1. found 464.2.

The starting material,(R)-2-bromo-1-(3-formamido-4-benzyloxyphenyl)ethanol, can be prepared asdescribed in U.S. Pat. No. 6,268,533 B1; or R. Hett et al., OrganicProcess Research and Development, 1998, 2:96-99; or using proceduressimilar to those described in Hong et al., Tetrahedron Lett., 1994,35:6631; or similar to those described in U.S. Pat. No. 5,495,054.

Preparation 872-((S)-{9-[(R)-2-(4-Benzyloxy-3-formylaminophenyl)-2-(tert-butyldimethyl-silanyloxy)ethylamino]nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

The product of Preparation 86 (500 mg, 1.008 mmol) and sodium iodide(243 mg, 1.62 mmol) are stirred in tetrahydrofuran (0.5 mL) for 15 minat ambient temperature. The product of Preparation 10 (1.29 mmol) andsodium bicarbonate (272 mg, 3.24 mmol) are then added and the reactionmixture is heated at 80° C. for 24 h. The reaction mixture is thenallowed to cool. Water (2 mL) is then added and the mixture is extractedwith dichloromethane (2×2 mL). The combined organic extracts are washedwith 1 M hydrochloric acid (2×1 mL), dried (magnesium sulfate) and thesolvent is removed under reduced pressure. The crude residue is purifiedby flash chromatography to give the title compound.

Preparation 882-((S)-1-{9-[(R)-2-(4-Benzyloxy-3-formylaminophenyl)-2-hydroxyethylamino]nonyl}-pyrrolidin-3-yl)-2,2-diphenylacetamide

To a stirred solution of the product of Preparation 87 (0.44 mmol) intetrahydrofuran (2.2 mL) at ambient temperature is added triethylaminetrihydrofluoride (108 μL, 0.66 mmol). The reaction mixture is stirredfor 24 h and then diluted with dichloromethane (5 mL) and washed with 1M hydrochloric acid (2 mL) and saturated aqueous sodium bicarbonate (2mL). The organic phase is dried (magnesium sulfate) and the solvent isremoved under reduced pressure to give the title compound.

Example 1512-((S)-1-{9-[(R)-2-(3-Formylamino-4-hydroxyphenyl)-2-hydroxyethylamino]nonyl}-pyrrolidin-3-yl)-2,2-diphenylacetamide

Palladium (10 wt. % (dry basis) on activated carbon) (124 mg) is addedto a stirred solution of the product of Preparation 88 (0.44 mmol) inethanol (4 mL) and the reaction mixture is placed under an atmosphere ofhydrogen. After stirring for 12 h, the reaction mixture is filteredthrough a pad of Celite, washed with methanol (2 mL) and the solvent isremoved under reduced pressure. The resulting residue is purified bypreparative HPLC to give the title compound as the ditrifluoroacetatesalt.

Examples 152-300

Using the methods described above and the appropriate startingmaterials, the compounds of Examples 152 to 300 as shown in Table II areprepared.

Preparation 896-(2-Bromo-(R)-1-tert-butyldimethylsilyloxy)ethyl-2,2-dimethyl-1,3-benzodioxan

(a) 6-Bromo-2,2-dimethyl-4H-benzo[1,3]dioxine

To 5-bromo-2-hydroxybenzyl alcohol (93 g, 0.46 mol, available fromSigma-Aldrich) in 2.0 L of 2,2-dimethoxypropane was added 700 mL ofacetone, followed by zinc chloride (170 g). After stirring for 18 hours,1.0 M aqueous sodium hydroxide was added until the aqueous phase wasbasic. Diethyl ether (1.5 L) was added to the slurry and the organicphase was decanted into a separatory funnel. The organic phase waswashed with brine, dried over Na₂SO₄, filtered and concentrated underreduced pressure to give the title compound as an oil.

(b) 6-Acetyl-2,2-dimethyl-4H-benzo[1,3]dioxine

To the product of step (a) (110 g, 0.46 mol) in 1.0 L of THF at −78° C.was added 236 mL (0.51 mol) of 2.14 M n-butyllithium in hexanes via adropping funnel. After 30 minutes, N-methyl-N-methoxy acetamide (71 g,0.69 mol, available from TCI) was added. After 2 hours, the reactionmixture was quenched with water, diluted with 2.0 L of 1.0 M aqueousphosphate buffer (pH=7.0) and extracted once with diethyl ether. Thediethyl ether phase was washed once with brine, dried over Na₂SO₄,filtered and concentrated under reduced pressure to give a light orangeoil. The oil was dissolved in a minimum volume of ethyl acetate, dilutedwith hexanes, and to give the title compound as a crystalline solid.

(c) 6-Bromoacetyl-2,2-dimethyl-4H-benzo[1,3]dioxine

To the product of step (b) (23.4 g, 0.113 mol) in 600 mL of THF at −78°C. was added 135 mL of 1.0 M sodium hexamethyldisilazane in THF(Sigma-Aldrich). After 1 hour, trimethylsilyl chloride (15.8 mL, 0.124mol) was added. After another 30 minutes, bromine (5.82 mL, 0.113 mol)was added. After 10 minutes, the reaction was quenched by diluting thereaction mixture with diethyl ether and pouring it onto 500 mL of 5%aqueous Na₂SO₃ premixed with 500 mL of 5% aqueous NaHCO₃. The phaseswere separated and the organic phase was washed with brine, dried overNa₂SO₄, filtered and concentrated under reduced pressure to give thetitle compound as an oil that solidified upon storage in the freezer.

(d) (R)-2-Bromo-1-(2,2-dimethyl-4H-benzo[1,3]dioxin-6-yl)ethanol

To the product of step (c) (10 g, 35.1 mmol) in 100 mL of THF was addedthe solid catalyst of Preparation 13, step (c)(1) (0.97 g, 3.5 mmol).The solution was cooled to between −20° C. and −10° C. and BH₃-THF (35mL, 35 mmol) diluted with 50 mL THF was added dropwise via a droppingfunnel. After the addition was complete, the reaction mixture wasallowed to warm to ambient temperature. After 30 minutes, the reactionmixture was quenched by slow addition of 50 mL of methanol and thenconcentrated to a thick oil. The oil was purified by silica gelchromatography eluted with 1:2 ethyl acetate/hexanes. The fractions werecombined and concentrated to give the title compound as an off-whitesolid.

(e)[(R)-2-Bromo-1-(2,2-dimethyl-4H-benzo[1,3]dioxin-6-yl)ethoxy]-tert-butyldimethylsilane

To the product of step (d) (10 g, 34.8 mmol) and imidazole (4.7 g, 69.7mmol) dissolved in 100 mL DMF was added tert-butyldimethylsilyl chloride(5.78 g, 38.3 mmol). The reaction mixture was stirred for 18 hours. Thereaction mixture was then partitioned between 200 mL of saturated sodiumchloride and 200 mL of diethyl ether. The aqueous layer was extractedwith 200 mL of diethyl ether. The organic layers were then combined,washed with saturated sodium chloride (3×100 mL), dried over MgSO₄ andconcentrated. The product was purified by silica gel chromatography,eluting with hexanes followed by 5% ethyl acetate in hexanes. Thedesired fractions were combined and concentrated to give the titlecompound as an oil.

Preparation 902-((S)-1-{9-[(R)-2-(tert-Butyldimethylsilanyloxy)-2-(2,2-dimethyl-4H-benzol-[1,3]dioxin-6-yl)ethylamino]nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

The product of Preparation 89 (802 mg, 2.00 mmol) and sodium iodide (300mg, 2.00 mmol) are stirred in tetrahydrofuran (0.77 mL) for 15 min atambient temperature. The product of Preparation 10 (1.54 mmol) andsodium bicarbonate (388 mg, 4.62 mmol) are added and the reactionmixture is heated at 80° C. for 24 h. The reaction mixture is thencooled and water (2 mL) is added. The mixture is then extracted withdichloromethane (2×2 mL). The combined organic extracts are dried(magnesium sulfate) and the solvent is removed under reduced pressure.The crude residue is purified by flash chromatography to give the titlecompound.

Preparation 912-((S)-1-{9-[(R)-2-(2,2-Dimethyl-4H-benzol-[1,3]dioxin-6-yl)-2-hydroxyethylamino]-nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

Triethylamine trihydrofluoride (342 μL, 2.10 mmol) is added to a stirredsolution of the Product of Preparation 90 (1.05 mmol) in dichloromethane(10.5 mL) at ambient temperature. The reaction mixture is stirred for 24h and it is then diluted with dichloromethane (20 mL) and washed withsaturated aqueous sodium bicarbonate (15 mL). The organic layer is dried(magnesium sulfate) and the solvent is removed under reduced pressure togive the title compound.

Example 3012-((S)-1-{9-[(R)-2-Hydroxy-2-(4-hydroxy-3-hydroxymethylphenyl)ethylamino]-nonyl}pyrrolidin-3-yl)-2,2-diphenylacetamide

Trifluoroacetic acid (2.80 mL) is added to a stirred solution of theproduct of Preparation 91 (0.93 mmol) in THF/H₂O (14 mL, 1:1) and thereaction mixture is stirred for 2 h at ambient temperature. The reactionmixture is concentrated under reduced pressure and dissolved in 20%MeCN/H₂O then purified by preparative HPLC to yield the title compound.

Preparation 92 2,2-Diphenyl-2-piperidin-4-ylacetamide

The title compound is prepared as the fumaric acid salt by the procedureof Walsh et al., J. Med. Chem. 1989, 105-118 (Methods P-S on page 115);or by the procedure in EP 0 178 947, published Apr. 23, 1986(Preparation 15 on page 19, lines 20-36).

Preparation 93 2-Cyclobutyl-2-phenyl-2-(S)-pyrrolidin-3-ylacetamide

The title compound is prepared by the procedure described in Preparation1 by substituting 2-cyclobutyl-2-phenylacetonitrile in place ofdiphenylacetonitrile. Alternatively, the reaction conditions describedin Examples 1 and 6 of U.S. Pat. No. 3,984,557, issued Oct. 5, 1976, maybe used by substituting 2-cyclobutyl-2-phenylacetonitrile in place ofa-phenyl-a-(2-pyridyl)acetonitrile, and toluene-4-sulfonic acid(S)-1-benzylpyrrolidin-3-yl ester in place of1-isobutyl-3-chloropyrrolidine.

Preparation 94 2-Cyclopentyl-2-phenyl-2-(S)-pyrrolidin-3-ylacetamide

The title compound is prepared by the procedure described in Preparation1 by substituting 2-cyclopentyl-2-phenylacetonitrile in place ofdiphenylacetonitrile. Alternatively, the reaction conditions describedin Examples 1 and 6 of U.S. Pat. No. 3,984,557, issued Oct. 5, 1976, maybe used by substituting 2-cyclopentyl-2-phenylacetonitrile in place ofa-phenyl-a-(2-pyridyl)acetonitrile, and toluene-4-sulfonic acid(S)-1-benzylpyrrolidin-3-yl ester in place of1-isobutyl-3-chloropyrrolidine.

Preparation 95 2-Cyclohexyl-2-phenyl-2-(S)-pyrrolidin-3-ylacetamide

The title compound is prepared by the procedure described in Preparation1 by substituting 2-cyclohexyl-2-phenylacetonitrile in place ofdiphenylacetonitrile. Alternatively, the reaction conditions describedin Examples 1 and 6 of U.S. Pat. No. 3,984,557, issued Oct. 5, 1976, maybe used by substituting 2-cyclohexyl-2-phenylacetonitrile in place ofa-phenyl-a-(2-pyridyl)acetonitrile, and toluene-4-sulfonic acid(S)-1-benzylpyrrolidin-3-yl ester in place of1-isobutyl-3-chloropyrrolidine.

Preparation 96 2-Phenyl-2-(pyridin-2-yl)-2-(S)-pyrrolidin-3-ylacetamide

The title compound is prepared by the procedures described in Examples 1and 6 of U.S. Pat. No. 3,984,557, issued Oct. 5, 1976, and using steps(c) and (d) of Preparation 1.

Preparation 97 2-Phenyl-2-(S)-pyrrolidin-3-yl-2-(thien-2-yl)acetamide

The title compound is prepared by the procedure described in Preparation1 by substituting 2-(thien-2-yl)-2-phenylacetonitrile in place ofdiphenylacetonitrile. Alternatively, the reaction conditions describedin Examples 1 and 6 of U.S. Pat. No. 3,984,557, issued Oct. 5, 1976, maybe used by substituting 2-(thien-2-yl)-2-phenylacetonitrile in place ofa-phenyl-a-(2-pyridyl)acetonitrile, and toluene-4-sulfonic acid(S)-1-benzylpyrrolidin-3-yl ester in place of1-isobutyl-3-chloropyrrolidine.

Preparation 98 2-(S)-Pyrrolidin-3-yl-2,2-di(thien-2-yl)acetamide

The title compound is prepared by the procedure described in Preparation1 by substituting dithien-2-yl)acetonitrile in place ofdiphenylacetonitrile. Alternatively, the reaction conditions describedin Examples 1 and 6 of U.S. Pat. No. 3,984,557, issued Oct. 5, 1976, maybe used by substituting di(thien-2-yl)acetonitrile in place ofa-phenyl-a-(2-pyridyl)acetonitrile, and toluene-4-sulfonic acid(S)-1-benzylpyrrolidin-3-yl ester in place of1-isobutyl-3-chloropyrrolidine.

Preparation 99 N,N-Dimethyl-2,2-diphenyl-2-pyrrolidin-3-ylacetamide

The title compound is prepared as the fumarate salt by the proceduredescribed in Example 5 of U.S. Pat. No. 4,002,766, issued Jan. 11, 1977.The fumarate salt of the title compound has a reported melting point of159-160° C.

Preparation 100 3-Phenyl-3-(S)-pyrrolidin-3-yl-1,3-dihydroindol-2-one

The title compound is prepared as described in WO 98/54167, published onDec. 5, 2000.

Preparation A Cell Culture and Membrane Preparation From CellsExpressing Human β₁, β₂ or β₃ Adrenergic Receptors

Chinese hamster ovarian (CHO) cell lines stably expressing cloned humanβ₁, β₂ or β₃ adrenergic receptors, respectively, were grown to near confluency in Hams F-12 media with 10% FBS in the presence of 500 μg/mLGeneticin. The cell monolayer was lifted with 2 mM EDTA in PBS. Cellswere pelleted by centrifugation at 1,000 rpm, and cell pellets wereeither stored frozen at −80° C. or membranes were prepared immediatelyfor use. For preparation of β₁ and β₂ receptor expressing membranes,cell pellets were re-suspended in lysis buffer (10 mM HEPES/HCl, 10 mMEDTA, pH 7.4 at 4° C.) and homogenized using a tight-fitting Dounceglass homogenizer (30 strokes) on ice. For the more protease-sensitiveβ₃ receptor expressing membranes, cell pellets were homogenated in lysisbuffer (10 mM Tris/HCl, pH 7.4) supplemented with one tablet of“Complete Protease Inhibitor Cocktail Tablets with 2 mM EDTA” per 50 mLbuffer (Roche Catalog No. 1697498, Roche Molecular Biochemicals,Indianapolis, Ind.). The homogenate was centrifuged at 20,000×g, and theresulting pellet was washed once with lysis buffer by re-suspension andcentrifugation as above. The final pellet was then re-suspended inice-cold binding assay buffer (75 mM Tris/HCl pH 7.4, 12.5 mM MgCl₂, 1mM EDTA). The protein concentration of the membrane suspension wasdetermined by the methods described in Lowry et al., 1951, Journal ofBiological Chemistry, 193, 265; and Bradford, Analytical Biochemistry,1976, 72, 248-54. All membranes were stored frozen in aliquots at −80°C. or used immediately.

Preparation B Cell Culture and Membrane Preparation From CellsExpressing Human M₁, M₂, M₃ and M₄ Muscarinic Receptors

CHO cell lines stably expressing cloned human hM₁, hM₂, hM₃ and hM₄muscarinic receptor subtypes, respectively, were grown to nearconfluency in HAM's F-12 media supplemented with 10% FBS and 250 μg/mLGeneticin. The cells were grown in a 5% CO₂, 37° C. incubator and liftedwith 2 mM EDTA in dPBS. Cells were collected by 5 minute centrifugationat 650×g, and cell pellets were either stored frozen at −80° C. ormembranes were prepared immediately for use. For membrane preparation,cell pellets were resuspended in lysis buffer and homogenized with aPolytron PT-2100 tissue disrupter (Kinematica AG; 20 seconds×2 bursts).Crude membranes were centrifuged at 40,000×g for 15 minutes at 4° C. Themembrane pellet was then resuspended with re-suspension buffer andhomogenized again with the Polytron tissue disrupter. The proteinconcentration of the membrane suspension was determined by the methoddescribed in Lowry et al., 1951, Journal of Biochemistry, 193, 265. Allmembranes were stored frozen in aliquots at −80° C. or used immediately.Aliquots of prepared hM₅ receptor membranes were purchased directly fromPerkin Elmer and stored at −80° C. until use.

Assay Test Procedure A Radioligand Binding Assay for Human β₁, β₂ and β₃Adrenergic Receptors

Binding assays were performed in 96-well microtiter plates in a totalassay volume of 100 μL with 10-15 μg of membrane protein containing thehuman β₁, β₂ or β₃ adrenergic receptors in assay buffer (75 mM Tris/HClpH 7.4 at 25° C., 12.5 mM MgCl₂, 1 mM EDTA, 0.2% BSA). Saturationbinding studies for determination of K_(d) values of the radioligandwere done using [³H]-dihydroalprenolol (NET-720, 100 Ci/mmol,PerkinElmer Life Sciences Inc., Boston, Mass.) for the β₁, and β2receptors and [¹²⁵I]-(−)-iodocyanopindolol (NEX-189, 220 Ci/mmol,PerkinElmer Life Sciences Inc., Boston, Mass.) at 10 or 11 differentconcentrations ranging from 0.011 nM to 20 nM. Displacement assays fordetermination of K_(i) values of test compounds were done with[³H]-dihydroalprenolol at 1 nM and [¹²⁵I]-(−)-iodocyanopindolol at 0.5nM for 10 or 11 different concentrations of test compound ranging from10 μM to 10 μM. Non-specific binding was determined in the presence of10 μM propranolol. Assays were incubated for 1 hour at 37° C., and thenbinding reactions were terminated by rapid filtration over GF/B for theβ₁ and β₂ receptors or GF/C glass fiber filter plates for the β₃receptors (Packard BioScience Co., Meriden, Conn.) presoaked in 0.3%polyethyleneimine. Filter plates were washed three times with filtrationbuffer (75 mM Tris/HCl pH 7.4 at 4° C., 12.5 mM MgCl₂, 1 mM EDTA) toremove unbound radioactivity. The plates were then dried and 50 μL ofMicroscint-20 liquid scintillation fluid (Packard BioScience Co.,Meriden, Conn.) was added and plates were counted in a Packard Topcountliquid scintillation counter (Packard BioScience Co., Meriden, Conn.).Binding data were analyzed by nonlinear regression analysis with theGraphPad Prism Software package (GraphPad Software, Inc., San Diego,Calif.) using the 3-parameter model for one-site competition. The curveminimum was fixed to the value for nonspecific binding, as determined inthe presence of 10 μM propranolol. K_(i) values for test compounds werecalculated from observed IC₅₀ values and the K_(d) value of theradioligand using the Cheng-Prusoff equation (Cheng Y, and Prusoff W H.,Biochemical Pharmacology, 1973, 22, 23, 3099-108).

In this assay, a lower K_(i) value indicates that a test compound has ahigher binding affinity for the receptor tested. Exemplified compound ofthis invention that were tested in this assay typically were found tohave a K_(i) value of less than about 300 nM for the β₂ adrenergicreceptor. For example, the compound of Example 1 was found to have K_(i)value of less than 50 mM.

If desired, the receptor subtype selectivity for a test compound can becalculated as the ratio of K_(i)(β₁)/K_(i)(β₂) or K_(i)(β₃)/K_(i)(β₂).Typically, compounds of this invention demonstrated greater binding atthe β₂ adrenergic receptor compared to the β₁ or β₃ adrenergic receptor,i.e. K_(i)(β₁) or K_(i)(β₃) is typically greater than K_(i)(β₂).Generally, compounds having selectivity for the β₂ adrenergic receptorover the β₁ or β₃ adrenergic receptors are preferred; especiallycompounds having a selectivity greater than about 5; and in particular,greater than about 8. By way of example, the compound of Example 1 had aratio of K_(i)(β₁)/K_(i)(β₂) greater than 8.

Assay Test Procedure B Radioligand Binding Assay for MuscarinicReceptors

Radioligand binding assays for cloned human muscarinic receptors wereperformed in 96-well microtiter plates in a total assay volume of 100μL. CHO cell membranes stably expressing either the hM₁, hM₂, hM₃, hM₄or hM₅ muscarinic subtype were diluted in assay buffer to the followingspecific target protein concentrations (μg/well): 10 μg for hM₁, 10-15μg for hM₂, 10-20 μg for hM₃, 10-20 μg for hM₄, and 10-12 μg for hM₅ toget similar signals (cpm). The membranes were briefly homogenized usinga Polytron tissue disrupter (10 seconds) prior to assay plate addition.Saturation binding studies for determining K_(D) values of theradioligand were performed using L-[N-methyl-³H]scopolamine methylchloride ([³H]-NMS) (TRK666, 84.0 Ci/mmol, Amersham Pharmacia Biotech,Buckinghamshire, England) at concentrations ranging from 0.001 nM to 20nM. Displacement assays for determination of K_(i) values of testcompounds were performed with [³H]-NMS at 1 nM and eleven different testcompound concentrations. The test compounds were initially dissolved toa concentration of 400 μM in dilution buffer and then serially diluted5× with dilution buffer to final concentrations ranging from 10 μM to100 μM. The addition order and volumes to the assay plates were asfollows: 25 μL radioligand, 25 μL diluted test compound, and 50 μLmembranes. Assay plates were incubated for 60 minutes at 37° C. Bindingreactions were terminated by rapid filtration over GF/B glass fiberfilter plates (PerkinElmer Inc., Wellesley, Mass.) pre-treated in 1%BSA. Filter plates were rinsed three times with wash buffer (10 mMHEPES) to remove unbound radioactivity. The plates were then air driedand 50 μL Microscint-20 liquid scintillation fluid (PerkinElmer Inc.,Wellesley, Mass.) was added to each well. The plates were then countedin a PerkinElmer Topcount liquid scintillation counter (PerkinElmerInc., Wellesley, Mass.). Binding data were analyzed by nonlinearregression analysis with the GraphPad Prism Software package (GraphPadSoftware, Inc., San Diego, Calif.) using the one-site competition model.K_(i) values for test compounds were calculated from observed IC₅₀values and the K_(D) value of the radioligand using the Cheng-Prusoffequation (Cheng Y; Prusoff W H. (1973) Biochemical Pharmacology,22(23):3099-108). K_(i) values were converted to pK_(i) values todetermine the geometric mean and 95% confidence intervals. These summarystatistics were then converted back to K_(i) values for data reporting.

In this assay, a lower K_(i) value indicates that the test compound hasa higher binding affinity for the receptor tested. Exemplified compoundof this invention that were tested in this assay typically were found tohave a K_(i) value of less than about 300 nM for the M₃ muscarinicreceptor. For example, the compound of Example 1 was found to have K_(i)values of less than 50 nM.

Assay Test Procedure C Whole-cell cAMP Flashplate Assay in CHO CellLines Heterologously Expressing Human β₁, β₂ or β₃ Adrenergic Receptors

cAMP assays were performed in a radioimmunoassay format using theFlashplate Adenylyl Cyclase Activation Assay System with [¹²⁵I]-cAMP(NEN SMP004, PerkinElmer Life Sciences Inc., Boston, Mass.), accordingto the manufacturers instructions. For the determination of β receptoragonist potency (EC₅₀), CHO-K1 cell lines stably expressing cloned humanβ₁, β₂ or β₃ adrenergic receptors were grown to near confluency in HAM'sF-12 media supplemented with 10% FBS and Geneticin (250 μg/mL). Cellswere rinsed with PBS and detached in dPBS (Dulbecco's Phosphate BufferedSaline, without CaCl₂ and MgCl₂) containing 2 mM EDTA or Trypsin-EDTAsolution (0.05% trypsin/0.53 mM EDTA). After counting cells in Coultercell counter, cells were pelleted by centrifugation at 1,000 rpm andre-suspended in stimulation buffer containing IBMX (PerkinElmer Kit)pre-warmed to room temperature to a concentration of 1.6×10⁶ to 2.8×10⁶cells/mL. About 60,000 to 80,000 cells per well were used in this assay.Test compounds (10 mM in DMSO) were diluted into PBS containing 0.1% BSAin Beckman Biomek-2000 and tested at 11 different concentrations rangingfrom 100 μM to 1 μM. Reactions were incubated for 10 min at 37° C. andstopped by adding 100 μL of cold detection buffer containing [¹²⁵I]-cAMP(NEN SMP004, PerkinElmer Life Sciences, Boston, Mass.). The amount ofcAMP produced (pmol/well) was calculated based on the counts observedfor the samples and cAMP standards as described in the manufacturer'suser manual. Data were analyzed by nonlinear regression analysis withthe GraphPad Prism Software package (GraphPad Software, Inc., San Diego,Calif.) with the sigmoidal equation. The Cheng-Prusoff equation (ChengY, and Prusoff W H., Biochemical Pharmacology, 1973, 22, 23, 3099-108)was used to calculate the EC50 values.

In this assay, a lower EC₅₀ value indicates that the test compound has ahigher functional activity at the receptor tested. Exemplified compoundof this invention that were tested in this assay typically were found tohave a EC₅₀ value of less than about 300 nM for the β₂ adrenergicreceptor. For example, the compound of Example 1 was found to have EC₅₀values of less than 10 nM.

If desired, the receptor subtype selectivity for a test compound can becalculated as the ratio of EC₅₀(β₁)/EC₅₀(β₂) or EC₅₀(β₃)/EC₅₀(β₂).Typically, compounds of this invention demonstrated greater functionalactivity at the β₂ adrenergic receptor compared to the β₁ or β₃adrenergic receptor, i.e. EC₅₀(β₁) or EC₅₀(β₃) is typically greater thanEC₅₀(β₂). Generally, compounds having selectivity for the β2 adrenergicreceptor over the β₁ or β₃ adrenergic receptors are preferred;especially compounds having a selectivity greater than about 5; and inparticular, greater than about 10. By way of example, the compound ofExample 1 had ratios of EC₅₀(β₁)/EC₅₀(β₂) greater than 10.

Assay Test Procedure D Functional Assays of Antagonism for MuscarinicReceptor Subtypes

A. Blockade of Agonist-Mediated Inhibition of cAMP Accumulation

In this assay, the functional potency of a test compound is determinedby measuring the ability of the test compound to blockoxotremorine-inhibition of forskolin-mediated cAMP accumulation inCHO-K1 cells expressing the hM₂ receptor. cAMP assays are performed in aradioimmunoassay format using the Flashplate Adenylyl Cyclase ActivationAssay System with ¹²⁵I-cAMP (NEN SMP004B, PerkinElmer Life SciencesInc., Boston, Mass.), according to the manufacturer's instructions.Cells are rinsed once with dPBS and lifted with Trypsin-EDTA solution(0.05% trypsin/0.53 mM EDTA) as described in the Cell Culture andMembrane Preparation section above. The detached cells are washed twiceby centrifugation at 650×g for five minutes in 50 mL dPBS. The cellpellet is then re-suspended in 10 mL dPBS, and the cells are countedwith a Coulter Z1 Dual Particle Counter (Beckman Coulter, Fullerton,Calif.). The cells are centrifuged again at 650×g for five minutes andre-suspended in stimulation buffer to an assay concentration of1.6×10⁶-2.8×10⁶ cells/mL.

The test compound is initially dissolved to a concentration of 400 μM indilution buffer (dPBS supplemented with 1 mg/mL BSA (0.1%)), and thenserially diluted with dilution buffer to final molar concentrationsranging from 100 μM to 0.1 nM. Oxotremorine is diluted in a similarmanner.

To measure oxotremorine inhibition of adenylyl cyclase (AC) activity, 25μL forskolin (25 μM final concentration diluted in dPBS), 25 μL dilutedoxotremorine, and 50 μL cells are added to agonist assay wells. Tomeasure the ability of a test compound to block oxotremorine-inhibitedAC activity, 25 μL forskolin and oxotremorine (25 μM and 5 μM finalconcentrations, respectively, diluted in dPBS), 25 μL diluted testcompound, and 50 μL cells are added to remaining assay wells.

Reactions are incubated for 10 minutes at 37° C. and stopped by additionof 100 μL ice-cold detection buffer. Plates are sealed, incubatedovernight at room temperature and counted the next morning on aPerkinElmer TopCount liquid scintillation counter (PerkinElmer Inc.,Wellesley, Mass.). The amount of cAMP produced (μmol/well) is calculatedbased on the counts observed for the samples and cAMP standards, asdescribed in the manufacturer's user manual. Data is analyzed bynonlinear regression analysis with the GraphPad Prism Software package(GraphPad Software, Inc., San Diego, Calif.) using the non-linearregression, one-site competition equation. The Cheng-Prusoff equation isused to calculate the K_(i), using the EC₅₀ of the oxotremorineconcentration-response curve and the oxotremorine assay concentration asthe K_(D) and [L], respectively.

In this assay, a lower K_(i) value indicates that the test compound hasa higher functional activity at the receptor tested. Exemplifiedcompound of this invention are expected to have a K_(i) value of lessthan about 300 nM for blockade of oxotremorine-inhibition offorskolin-mediated cAMP accumulation in CHO-K1 cells expressing the hM₂receptor.

B. Blockade of Agonist-Mediated [³⁵S]GTPγS Binding

In a second functional assay, the functional potency of test compoundscan be determined by measuring the ability of the compounds to blockoxotremorine-stimulated [³⁵S]GTPγS_binding in CHO-K1 cells expressingthe hM₂ receptor.

At the time of use, frozen membranes are thawed and then diluted inassay buffer with a final target tissue concentration of 5-10 μg proteinper well. The membranes are briefly homogenized using a Polytron PT-2100tissue disrupter and then added to the assay plates.

The EC₉₀ value (effective concentration for 90% maximal response) forstimulation of [³⁵ S]GTPγS binding by the agonist oxotremorine isdetermined in each experiment.

To determine the ability of a test compound to inhibitoxotremorine-stimulated [³⁵S]GTPγS binding, the following is added toeach well of 96 well plates: 25 μL of assay buffer with [³⁵S]GTPγS (0.4nM), 25 μL of oxotremorine (EC₉₀) and GDP (3 uM), 25 μL of diluted testcompound and 25 μL CHO cell membranes expressing the hM₂ receptor. Theassay plates are then incubated at 37° C. for 60 minutes. The assayplates are filtered over 1% BSA-pretreated GF/B filters using aPerkinElmer 96-well harvester. The plates are rinsed with ice-cold washbuffer for 3×3 seconds and then air or vacuum dried. Microscint-20scintillation liquid (50 μL) is added to each well, and each plate issealed and radioactivity counted on a Topcounter (PerkinElmer). Data areanalyzed by nonlinear regression analysis with the GraphPad PrismSoftware package (GraphPad Software, Inc., San Diego, Calif.) using thenon-linear regression, one-site competition equation. The Cheng-Prusoffequation is used to calculate the K_(i), using the IC₅₀ values of theconcentration-response curve for the test compound and the oxotremorineconcentration in the assay as the K_(D) and [L], ligand concentration,respectively.

In this assay, a lower K_(i) value indicates that the test compound hasa higher functional activity at the receptor tested. Exemplifiedcompound of this invention are expected to have a K_(i) value of lessthan about 300 nM for blockade of oxotremorine-stimulated[³⁵S]GTPγS_binding in CHO-K1 cells expressing the hM₂ receptor.

C. Blockade of Agonist-Mediated Calcium Release via FLIPR Assays

Muscarinic receptor subtypes (M₁, M₃ and M₅ receptors), which couple toG_(q) proteins, activate the phospholipase C (PLC) pathway upon agonistbinding to the receptor. As a result, activated PLC hydrolyzesphosphatyl inositol diphosphate (PIP₂) to diacylglycerol (DAG) andphosphatidyl-1,4,5-triphosphate (IP₃), which in turn generates calciumrelease from intracellular stores, i.e., endoplasmic and sarcoplasmicreticulum. The FLIPR (Molecular Devices, Sunnyvale, Calif.) assaycapitalizes on this increase in intracellular calcium by using a calciumsensitive dye (Fluo-4AM, Molecular Probes, Eugene, Oreg.) thatfluoresces when free calcium binds. This fluorescence event is measuredin real time by the FLIPR, which detects the change in fluorescence froma monolayer of cells cloned with human M₁ and M₃, and chimpanzee M₅receptors. Antagonist potency can be determined by the ability ofantagonists to inhibit agonist-mediated increases in intracellularcalcium.

For FLIPR calcium stimulation assays, CHO cells stably expressing thehM₁, hM₃ and cM₅ receptors are seeded into 96-well FLIPR plates thenight before the assay is done. Seeded cells are washed twice byCellwash (MTX Labsystems, Inc.) with FLIPR buffer (10 mM HEPES, pH 7.4,2 mM calcium chloride, 2.5 mM probenecid in Hank's Buffered SaltSolution (HBSS) without calcium and magnesium) to remove growth mediaand leaving 50 μL/well of FLIPR buffer. The cells are then incubatedwith 50 μL/well of 4 μM FLUO-4AM (a 2× solution was made) for 40 minutesat 37° C., 5% carbon dioxide. Following the dye incubation period, cellsare washed two times with FLIPR buffer, leaving a final volume of 50μL/well.

To determine antagonist potency, the dose-dependent stimulation ofintracellular Ca²⁺ release for oxotremorine is first determined so thatantagonist potency can later be measured against oxotremorinestimulation at an EC₉₀ concentration. Cells are first incubated withcompound dilution buffer for 20 minutes, followed by agonist addition,which is performed by the FLIPR. An EC₉₀ value for oxotremorine isgenerated according to the method detailed in the FLIPR measurement anddata reduction section below, in conjunction with the formulaEC_(F)=((F/100-F)^1/H)*EC₅₀. An oxotremorine concentration of 3×EC_(F)is prepared in stimulation plates such that an EC₉₀ concentration ofoxotremorine is added to each well in the antagonist inhibition assayplates.

The parameters used for the FLIPR are: exposure length of 0.4 seconds,laser strength of 0.5 watts, excitation wavelength of 488 nm, andemission wavelength of 550 nm. Baseline is determined by measuring thechange in fluorescence for 10 seconds prior to addition of agonist.Following agonist stimulation, the FLIPR continuously measured thechange of fluorescence every 0.5 to 1 second for 1.5 minutes to capturethe maximum fluorescence change.

The change of fluorescence is expressed as maximum fluorescence minusbaseline fluorescence for each well. The raw data is analyzed againstthe logarithm of drug concentration by nonlinear regression withGraphPad Prism (GraphPad Software, Inc., San Diego, Calif.) using thebuilt-in model for sigmoidal dose-response. Antagonist K_(i) values aredetermined by Prism using the oxotremorine EC₅₀ value as the K_(D) andthe oxotremorine EC₉₀ for the ligand concentration according to theCheng-Prusoff equation (Cheng & Prusoff, 1973).

In this assay, a lower K_(i) value indicates that the test compound hasa higher functional activity at the receptor tested. Exemplifiedcompound of this invention are expected to have a K_(i) value of lessthan about 300 nM for blockade of agonist-mediated calcium release inCHO cells stably expressing the hM₁, hM₃ and cM₅ receptors.

Assay Test Procedure E Whole-cell cAMP Flashplate Assay With a LungEpithelial Cell Line Endogenously Expressing Human β₂ AdrenergicReceptor

For the determination of agonist potencies and efficacies (intrinsicactivities) in a cell line expressing endogenous levels of the β₂adrenergic receptor, a human lung epithelial cell line (BEAS-2B) wasused (ATCC CRL-9609, American Type Culture Collection, Manassas, Va.)(January B, et al., British Journal of Pharmacology, 1998, 123, 4,701-11). Cells were grown to 75-90% confluency in complete, serum-freemedium (LHC-9 MEDIUM containing Epinephrine and Retinoic Acid, cat#181-500, Biosource International, Camarillo, Calif.). The day beforethe assay, medium was switched to LHC-8 (no epinephrine or retinoicacid, cat #141-500, Biosource International, Camarillo, Calif.). cAMPassays were performed in a radioimmunoassay format using the FlashplateAdenylyl Cyclase Activation Assay System with [¹²⁵I]-cAMP (NEN SMP004,PerkinElmer Life Sciences Inc., Boston, Mass.), according to themanufacturers instructions. On the day of the assay, cells were rinsedwith PBS, lifted by scraping with 5 mM EDTA in PBS, and counted. Cellswere pelleted by centrifugation at 1,000 rpm and re-suspended instimulation buffer pre-warmed to 37° C. at a final concentration of600,000 cells/mL. Cells were used at a final concentration of 100,000 to120,000 cells/well in this assay. Test compounds were serially dilutedinto assay buffer (75 mM Tris/HCl pH 7.4 at 25° C., 12.5 mM MgCl₂, 1 mMEDTA, 0.2% BSA) in Beckman Biomek-2000. Test compounds were tested inthe assay at 11 different concentrations, ranging from 10 μM to 10 pM.Reactions were incubated for 10 min at 37° C. and stopped by addition of100 μL of ice-cold detection buffer. Plates were sealed, incubated overnight at 4° C. and counted the next morning in a Topcount scintillationcounter (Packard BioScience Co., Meriden, Conn.). The amount of cAMPproduced per mL of reaction was calculated based on the counts observedfor samples and cAMP standards, as described in the manufacturer's usermanual. Data were analyzed by nonlinear regression analysis with theGraphPad Prism Software package (GraphPad Software, Inc., San Diego,Calif.) using the 4-parameter model for sigmoidal dose-response.

In this assay, a lower EC₅₀ value indicates that the test compound has ahigher functional activity at the receptor tested. Exemplified compoundof this invention that were tested in this assay typically were found tohave a EC₅₀ value of less than about 300 nM for the β₂ adrenergicreceptor. For example, the compound of Example 1 was found to have EC₅₀values of less than 10 nM.

If desired, test compound efficacy (% Eff) was calculated from the ratioof the observed Emax (TOP of the fitted curve) and the maximal responseobtained for isoproterenol dose response curve and was expressed as %Eff relative to isoproterenol. Exemplified compounds of this inventiontested in this assay typically demonstrated a % Eff greater than about40.

Assay Test Procedure F Duration of Bronchoprotection In Guinea PigModels of Acetylcholine-Induced or Histamine-Induced Bronchoconstriction

These in vivo assays are used to assess the bronchoprotective effects oftest compounds exhibiting both muscarinic receptor antagonist and β₂adrenergic receptor agonist activity. To isolate muscarinic antagonistactivity in the acetylcholine-induced bronchoconstriction model, theanimals are administered propanolol, a compound that blocks β receptoractivity, prior to the administration of acetylcholine. Duration ofbronchoprotection in the histamine-induced bronchoconstriction modelreflects β₂ adrenergic receptor agonist activity.

Groups of 6 male guinea pigs (Duncan-Hartley (HsdPoc:DH) Harlan,Madison, Wis.) weighing between 250 and 350 g are individuallyidentified by cage cards. Throughout the study, animals are allowedaccess to food and water ad libitum.

Test compounds are administered via inhalation over 10 minutes in awhole-body exposure dosing chamber (R&S Molds, San Carlos, Calif.). Thedosing chambers are arranged so that an aerosol is simultaneouslydelivered to 6 individual chambers from a central manifold. Guinea pigsare exposed to an aerosol of a test compound or vehicle (WFI). Theseaerosols are generated from aqueous solutions using an LC Star NebulizerSet (Model 22F51, PARI Respiratory Equipment, Inc. Midlothian, Va.)driven by a mixture of gases (CO₂=5%, O₂=21% and N₂=74%) at a pressureof 22 psi. The gas flow through the nebulizer at this operating pressureis approximately 3 L/minute. The generated aerosols are driven into thechambers by positive pressure. No dilution air is used during thedelivery of aerosolized solutions. During the 10 minute nebulization,approximately 1.8 mL of solution is nebulized. This value is measuredgravimetrically by comparing pre- and post-nebulization weights of thefilled nebulizer.

The bronchoprotective effects of test compounds administered viainhalation are evaluated using whole body plethysmography at 1.5, 24, 48and 72 hours post-dose.

Forty-five minutes prior to the start of the pulmonary evaluation, eachguinea pig is anesthetized with an intramuscular injection of ketamine(43.75 mg/kg), xylazine (3.50 mg/kg) and acepromazine (1.05 mg/kg).After the surgical site is shaved and cleaned with 70% alcohol, a 2-3 cmmidline incision of the ventral aspect of the neck is made. Then, thejugular vein is isolated and cannulated with a saline-filledpolyethylene catheter (PE-50, Becton Dickinson, Sparks, Md.) to allowfor intravenous infusions of acetylcholine (Ach) or histamine in saline.The trachea is then dissected free and cannulated with a 14G teflon tube(#NE-014, Small Parts, Miami Lakes, Fla.). If required, anesthesia ismaintained by additional intramuscular injections of the aforementionedanesthetic mixture. The depth of anesthesia is monitored and adjusted ifthe animal responds to pinching of its paw or if the respiration rate isgreater than 100 breaths/minute.

Once the cannulations are completed, the animal is placed into aplethysmograph (#PLY3114, Buxco Electronics, Inc., Sharon, Conn.) and anesophageal pressure cannula (PE-160, Becton Dickinson, Sparks, Md.) isinserted to measure pulmonary driving pressure (pressure). The teflontracheal tube is attached to the opening of the plethysmograph to allowthe guinea pig to breathe room air from outside the chamber. The chamberis then sealed. A heating lamp is used to maintain body temperature andthe guinea pig's lungs are inflated 3 times with 4 mL of air using a 10mL calibration syringe (#5520 Series, Hans Rudolph, Kansas City, Mo.) toensure that the lower airways did not collapse and that the animal didnot suffer from hyperventilation.

Once it was determined that baseline values are within the range of0.3-0.9 mL/cm H₂O for compliance and within the range of 0.1-0.199 cmH₂O/mL per second for resistance, the pulmonary evaluation is initiated.A Buxco pulmonary measurement computer program enabled the collectionand derivation of pulmonary values.

Starting this program initiated the experimental protocol and datacollection. The changes in volume over time that occur within theplethysmograph with each breath are measured via a Buxco pressuretransducer. By integrating this signal over time, a measurement of flowis calculated for each breath. This signal, together with the pulmonarydriving pressure changes, which are collected using a Sensym pressuretransducer (#TRD4100), is connected via a Buxco (MAX 2270) preamplifierto a data collection interface (#'s SFT3400 and SFT3813). All otherpulmonary parameters are derived from these two inputs.

Baseline values are collected for 5 minutes, after which time the guineapigs are challenged with Ach or histamine. When evaluating themuscarinic antagonist effects, propanolol (5 mg/Kg, iv) (Sigma-Aldrich,St. Louis, Mo.) is administered 15 minutes prior to challenge with Ach.Ach (Sigma-Aldrich, St. Louis, Mo.) (0.1 mg/mL) is infused intravenouslyfor 1 minute from a syringe pump (sp210 iw, World Precision Instruments,Inc., Sarasota, Fla.) at the following doses and prescribed times fromthe start of the experiment: 1.9 μg/minute at 5 minutes, 3.8 μg/minuteat 10 minutes, 7.5 μg/minute at 15 minutes, 15.0 μg/minute at 20minutes, 30 μg/minute at 25 minutes and 60 μg/minute at 30 minutes.Alternatively, bronchoprotection of test compounds is assessed in theacetylcholine challenge model without pretreatment with a beta blockingcompound.

When evaluating the β₂ adrenergic receptor agonist effects of testcompounds, histamine (25 μg/mL) (Sigma-Aldrich, St. Louis, Mo.) isinfused intravenously for 1 minute from a syringe pump at the followingdoses and prescribed times from the start of the experiment: 0.5μg/minute at 5 minutes, 0.9 μg/minute at 10 minutes, 1.9 μg/minute at 15minutes, 3.8 μg/minute at 20 minutes, 7.5 μg/minute at 25 minutes and 15μg/minute at 30 minutes. If resistance or compliance does not returnedto baseline values at 3 minutes following each Ach or histamine dose,the guinea pig's lungs are inflated 3 times with 4 mL of air from a 10mL calibration syringe. Recorded pulmonary parameters includerespiration frequency (breaths/minute), compliance (mL/cm H₂O) andpulmonary resistance (cm H₂O/mL per second). Once the pulmonary functionmeasurements are completed at minute 35 of this protocol, the guinea pigis removed from the plethysmograph and euthanized by carbon dioxideasphyxiation.

The data were evaluated in one of two ways:

(a) Pulmonary resistance (R_(L), cm H₂O/mL per second) is calculatedfrom the ratio of “change in pressure” to “the change in flow.” TheR_(L) response to ACh (60 μg/min, IH) is computed for the vehicle andthe test compound groups. The mean ACh response in vehicle-treatedanimals, at each pre-treatment time, is calculated and used to compute %inhibition of ACh response, at the corresponding pre-treatment time, ateach test compound dose. Inhibition dose-response curves for ‘R_(L)’were fitted with a four parameter logistic equation using GraphPadPrism, version 3.00 for Windows (GraphPad Software, San Diego, Calif.)to estimate bronchoprotective ID₅₀ (dose required to inhibit the ACh (60μg/min) bronchocontrictor response by 50%). The equation used is asfollows:Y=Min+(Max−Min)/(1+10^(((log ID50-X)*Hillslope)))where X is the logarithm of dose, Y is the response (% Inhibition of AChinduced increase in R_(L)). Y starts at Min and approachesasymptotically to Max with a sigmoidal shape.

(b) The quantity PD₂, which is defined as the amount of Ach or histamineneeded to cause a doubling of the baseline pulmonary resistance, iscalculated using the pulmonary resistance values derived from the flowand the pressure over a range of Ach or histamine challenges using thefollowing equation (derived from the equation used to calculate PC₂₀values in the clinic (see Am. Thoracic Soc, 2000):

${PD}_{2} = {{antilog}\left\lbrack {{\log\; C_{1}} + \frac{\left( {{\log\; C_{2}} - {\log\; C_{1}}} \right)\left( {{2R_{0}} - R_{1}} \right)}{R_{2} - R_{1}}} \right\rbrack}$

where:

-   -   C₁=concentration of Ach or histamine preceding C₂    -   C₂=concentration of Ach or histamine resulting in at least a        2-fold increase in pulmonary resistance (R_(L))    -   R₀=Baseline R_(L) value    -   R₁=R_(L) value after C₁    -   R₂=R_(L) value after C₂

Statistical analysis of the data is performed using a twotailed-Students t-test. A P-value<0.05 was considered significant.

Exemplified compounds of this invention are expected to produce adose-dependent bronchoprotective effect against MCh-inducedbronchoconstriction and His-induced bronchoconstriction. Test compoundshaving a potency (ID₅₀ at 1.5 h post-dose) of less than about 300 μg/mLfor ACh-induced bronchoconstriction and less than about 300 μg/mL forHis-induced bronchoconstriction in this assay are generally preferred.Additionally, test compounds having a duration (PD T_(1/2)) ofbrochoprotective activity of at least about 24 hours in this assay aregenerally preferred.

Assay Test Procedure G Einthoven Model for Measuring Changes inVentilation in Guinea Pigs

The bronchodilator activity of test compounds is evaluated in ananesthetized guinea pig model (the Einthoven model), which usesventilation pressure as a surrogate measure of airway resistance. See,for example, Einthoven (1892) Pfugers Arch. 51: 367-445; and Mohammed etal. (2000) Pulm Pharmacol Ther. 13(6):287-92. In this model, muscarinicantagonist and β₂ agonist activity is assessed by determining theprotective effects against methacholine (MCh) and histamine(His)-induced bronchoconstriction.

This assay is conducted using Duncan-Hartley guinea pigs (Harlan,Indianapolis, Ind.), weighing between 300 and 400 g.

The test compound or vehicle (i.e., sterile water) is dosed byinhalation (IH) over a 10 minute time period in a whole body exposuredosing chamber (R+S Molds, San Carlos, Calif.) using 5 mL of dosingsolution. Animals are exposed to an aerosol, which is generated from anLC Star Nebulizer Set (Model 22F51, PARI Respiratory Equipment, Inc.Midlothian, Va.) driven by Bioblend a mixture of gasses (5% Co₂; 21% O₂;and 74% N₂) at a pressure of 22 psi. Pulmonary function is evaluated atvarious time-points after inhalation dosing.

Forty five minutes prior to the start of pulmonary function evaluation,the guinea pigs are anesthetized with an intramuscular (IM) injection ofa mixture of ketamine (13.7 mg/kg/xylazine (3.5 mg/kg)/acepromazine(1.05 mg/kg). A supplemental dose of this mixture (50% of initial dose)is administered as needed. The jugular vein and carotid artery areisolated and cannulated with saline-filled polyethylene catheters(micro-renathane and PE-50, respectively, Beckton Dickinson, Sparks,Md.). The carotid artery is connected to a pressure transducer to allowthe measurement of blood pressure and the jugular vein cannula is usedfor IV injection of either MCh or His. The trachea is then dissectedfree and cannulated with a 14G needle (#NE-014, Small Parts, MiamiLakes, Fla.). Once the cannulations are complete, the guinea pigs areventilated using a respirator (Model 683, Harvard Apparatus, Inc., MA)set at a stroke volume of 1 mL/100 g body weight but not exceeding 2.5mL volume, and at a rate of 100 strokes per minute. Ventilation pressure(VP) is measured in the tracheal cannula using a Biopac transducer thatis connected to a Biopac (TSD 137C) pre-amplifier. Body temperature ismaintained at 37° C. using a heating pad. Prior to initiating datacollection, pentobarbital (25 mg/kg) is administered intraperitoneally(IP) to suppress spontaneous breathing and obtain a stable baseline. Thechanges in VP are recorded on a Biopac Windows data collectioninterface. Baseline values are collected for at least 5 minutes, afterwhich time guinea pigs are challenged IV non-cumulatively with 2-foldincremental doses of the bronchoconstrictor (MCh or His). When MCh isused as the bronchoconstrictor agent, animals are pre-treated withpropranolol (5 mg/kg, IV) to isolate the antimuscarinic effects of thetest compound. Changes in VP are recorded using the Acknowledge DataCollection Software (Santa Barbara, Calif.). After the completion ofstudy, the animals are euthanized.

Change in VP is measured in cm of water. Change in VP (cm H₂O)=peakpressure (after bronchoconstrictor challenge)−peak baseline pressure.The dose-response curve to MCh or His is fitted to a four parameterlogistic equation using GraphPad Prism, version 3.00 for Windows(GraphPad Software, San Diego, Calif.). The equation used is as follows:Y=Min+(Max−Min)/(1+10^(((log ID50-X)*Hillslope)))where X is the logarithm of dose, Y is the response. Y starts at Min andapproaches asymptotically to Max with a sigmoidal shape.

The percent inhibition of the bronchoconstrictor response to asubmaximal dose of MCh or His is calculated at each dose of the testcompound using the following equation: % Inhibition ofresponse=100−((peak pressure (after bronchoconstrictor challenge,treated)−peak baseline pressure (treated)*100%/(peak pressure (afterbronchoconstrictor challenge, water)−peak baseline pressure (water)).Inhibition curves are fitted using the four parameter logistic equationfrom GraphPad software. ID₅₀ (dose required to produce 50% inhibition ofthe bronchoconstrictor response) and Emax (maximal inhibition) are alsoestimated wherever appropriate.

The magnitude of bronchoprotection at different time-points afterinhalation of the test compound is used to estimate the pharmacodynamichalf-life (PD T_(1/2)). PD T_(1/2) is determined using a non-linearregression fit using a one-phase exponential decay equation (GraphPadPrism, Version 4.00): Y=Span*exp(−K*X)+Plateau; Starts at Span+Plateauand decays to Plateau with a rate constant K. The PD T_(1/2=0.69)/K.Plateau is constrained to 0.

Exemplified compounds of this invention are expected to produce adose-dependent bronchoprotective effect against MCh-inducedbronchoconstriction and His-induced bronchoconstriction. Generally, testcompounds having an ID₅₀ less than about 300 μg/mL for MCh-inducedbronchoconstriction and an ID₅₀ less than about 300 μg/mL forHis-induced bronchoconstriction at 1.5 hours post-dose in this assay arepreferred. Additionally, test compounds having a duration (PD T_(1/2))of brochoprotective activity of at least about 24 hours in this assayare generally preferred.

Assay Test Procedure H Inhalation Guinea Pig Salivation Assay

Guinea pigs (Charles River, Wilmington, Mass.) weighing 200-350 g areacclimated to the in-house guinea pig colony for at least 3 daysfollowing arrival. Test compound or vehicle are dosed via inhalation(IH) over a 10 minute time period in a pie shaped dosing chamber (R+SMolds, San Carlos, Calif.). Test solutions are dissolved in sterilewater and delivered using a nebulizer filled with 5.0 mL of dosingsolution. Guinea pigs are restrained in the inhalation chamber for 30minutes. During this time, guinea pigs are restricted to an area ofapproximately 110 sq. cm. This space is adequate for the animals to turnfreely, reposition themselves, and allow for grooming. Following 20minutes of acclimation, guinea pigs are exposed to an aerosol generatedfrom a LS Star Nebulizer Set (Model 22F51, PARI Respiratory Equipment,Inc. Midlothian, Va.) driven by house air at a pressure of 22 psi. Uponcompletion of nebulization, guinea pigs are evaluated at 1.5, 6, 12, 24,48, or 72 hrs after treatment.

Guinea pigs are anesthetized one hour before testing with anintramuscular (IM) injection of a mixture of ketamine 43.75 mg/kg,xylazine 3.5 mg/kg, and acepromazine 1.05 mg/kg at an 0.88 mL/kg volume.Animals are placed ventral side up on a heated (37° C.) blanket at a 20degree incline with their head in a downward slope. A 4-ply 2×2 inchgauze pad (Nu-Gauze General-use sponges, Johnson and Johnson, Arlington,Tex.) is inserted in the guinea pig's mouth. Five minutes later, themuscarinic agonist pilocarpine (3.0 mg/kg, s.c.) is administered and thegauze pad is immediately discarded and replaced by a new pre-weighedgauze pad. Saliva is collected for 10 minutes, at which point the gauzepad is weighed and the difference in weight recorded to determine theamount of accumulated saliva (in mg). The mean amount of salivacollected for animals receiving the vehicle and each dose of testcompound is calculated. The vehicle group mean is considered to be 100%salivation. Results are calculated using result means (n=3 or greater).Confidence intervals (95%) are calculated for each dose at each timepoint using two-way ANOVA. This model is a modified version of theprocedure described in Rechter, “Estimation of anticholinergic drugeffects in mice by antagonism against pilocarpine-induced salivation”Ata Pharmacol Toxicol, 1996, 24:243-254.

The mean weight of saliva in vehicle-treated animals, at eachpre-treatment time, is calculated and used to compute % inhibition ofsalivation, at the corresponding pre-treatment time, at each dose. Theinhibition dose-response data are fitted to a a four parameter logisticequation using GraphPad Prism, version 3.00 for Windows (GraphPadSoftware, San Diego, Calif.) to estimate anti-sialagogue ID₅₀ (doserequired to inhibit 50% of pilocarpine-evoked salivation). The equationused is as follows:Y=Min+(Max−Min)/(1+10^(((log ID50-X)*Hillslope)))where X is the logarithm of dose, Y is the response (% inhibition ofsalivation). Y starts at Min and approaches asymptotically to Max with asigmoidal shape.

The ratio of the anti-sialagogue ID₅₀ to bronchoprotective ID₅₀ is usedto compute the apparent lung-selectivity index of the test compound.Generally, compounds having an apparent lung-selectivity index greaterthan about 5 are preferred. In this assay, the compound of Example 3 hadan apparent lung-selectivity index greater than 5.

While the present invention has been described with reference tospecific aspects or embodiments thereof, it will be understood by thoseof ordinary skilled in the art that various changes can be made orequivalents can be substituted without departing from the true spiritand scope of the invention. Additionally, to the extent permitted byapplicable patent statues and regulations, all publications, patents andpatent applications cited herein are hereby incorporated by reference intheir entirety to the same extent as if each document had beenindividually incorporated by reference herein.

1. A compound of formula IV:

wherein R⁴ is a divalent group of the formula:—(R^(4a))_(d)-(A¹)_(e)-(R^(4b))_(f)-Q-(R^(4c))_(g)-(A²)_(h)-(R^(4d))_(i)—wherein d, e, f, g, h and i are each independently selected from 0 and1; R^(4a), R^(4b), R^(4c) and R^(4d) are each independently selectedfrom (1-10C)alkylene, (2-10C)alkenylene and (2-10C)alkynylene, whereineach alkylene, alkenylene or alkynylene group is unsubstituted orsubstituted with from 1 to 5 substituents independently selected from(1-4C)alkyl, fluoro, hydroxy, phenyl and phenyl-(1-4C)alkyl; A¹ and A²are each independently selected from (3-7C)cycloalkylene,(6-10C)arylene, —O-(6-10C)arylene, (6-10C)arylene-O—,(2-9C)heteroarylene, —O-(2-9C)heteroarylene, (2-9C)heteroarylene-O— and(3-6C)heterocyclene, wherein each cycloalkylene is unsubstituted orsubstituted with from 1 to 4 substituents selected independently from(1-4C)alkyl, and each arylene, heteroarylene or heterocyclene group isunsubstituted or substituted with from 1 to 4 substituents independentlyselected from halo, (1-4C)alkyl, (1-4C)alkoxy, —S-(1-4C)alkyl,—S(O)-(1-4C)alkyl, —S(O)₂-(1-4C)alkyl, —C(O)O(1-4C)alkyl, carboxy,cyano, hydroxy, nitro, trifluoromethyl and trifluoromethoxy; Q isselected from a bond, —O—, —C(O)O—, —OC(O)—, —S—, —S—(O)₂—,—N(Q^(a))C(O)—, —C(O)N(Q^(b))-, —N(Q^(c))S(O)₂—, —S(O)₂N(Q^(d))-,—N(Q^(e))C(O)N(Q^(f))-, —N(Q^(g))S(O)₂N(Q^(h))-, —OC(O)N(Q^(i))-,—N(Q^(j))C(O)O— and —N(Q^(k)); Q^(a), Q^(b), Q^(c), Q^(d), Q^(e), Q^(f),Q^(g), Q^(h), Q^(i), Q^(j) and Q^(k) are each independently selectedfrom hydrogen, (1-6C)alkyl, A³ and (1-4C)alkylene-A⁴, wherein the alkylgroup is unsubstituted or substituted with from 1 to 3 substituentsindependently selected from fluoro, hydroxy and (1-4C)alkoxy; ortogether with the nitrogen atom and the group R^(4b) or R^(4c) to whichthey are attached, form a 4-6 membered azacycloalkylene group; A³ and A⁴are each independently selected from (3-6C)cycloalkyl, (6-10C)aryl,(2-9C)heteroaryl and (3-6C)heterocyclyl, wherein each cycloalkyl isunsubstituted or substituted with from 1 to 4 substituents selectedindependently from (1-4C)alkyl and each aryl, heteroaryl or heterocyclylgroup is unsubstituted or substituted with from 1 to 4 substituentsindependently selected from halo, (1-4C)alkyl and (1-4C)alkoxy; providedthat the number of contiguous atoms in the shortest chain between thetwo nitrogen atoms to which R⁴ is attached is in the range of from 4 to16; or a pharmaceutically acceptable salt or stereoisomer thereof. 2.The compound of claim 1, wherein the number of contiguous atoms in theshortest chain between the two nitrogen atoms to which R⁴ is attached isin the range of from 8 to
 14. 3. The compound of claim 1, wherein thenumber of contiguous atoms in the shortest chain between the twonitrogen atoms to which R⁴ is attached is 8, 9, 10 or
 11. 4. Thecompound of claim 1, wherein R⁴ is a divalent group of the formula:—(R^(4a))_(d)— where R^(4a) is (4-10C)alkylene.
 5. The compound of claim4, wherein R⁴ is —(CH₂)₈—, —(CH₂)₉, and —(CH₂)₁₀—.
 6. The compound ofclaim 1, wherein R⁴ is a divalent group of the formula:—(R^(4a))_(d)-(A²)_(h)-(R^(4d))_(i)— wherein R^(4a) is (1-10C)alkylene;A² is (6-10C)arylene or (2-9C)heteroarylene; and R^(4d) is(1-10C)alkylene.
 7. The compound of claim 1, wherein R⁴ is a divalentgroup of the formula:—(R^(4a))_(d)-Q-(A²)_(h)-(R^(4d))_(i)— wherein Q is —O— or —N(Q^(k))-;Q^(k) is hydrogen or (1-3C)alkyl; R^(4a) is (1-10C)alkylene; A² is(6-10C)arylene or (2-9C)heteroarylene; and R^(4d) is (1-10C)alkylene. 8.The compound of claim 1, wherein Q is —N(Q^(a))C(O)— or —C(O)N(Q^(b))-.9. The compound of claim 8 wherein R⁴ is selected from:

wherein m is an integer from 2 to 10; and n is an integer from 2 to 10;provided that m+n is an integer from 4 to 12;

wherein o is an integer from 2 to 7; and p is an integer from 1 to 6;provided that o+p is an integer from 3 to 8; and wherein thephen-1,4-ylene group is optionally substituted with from 1 to 4substituents independently selected from halo, (1-4C)alkyl,(1-4C)alkoxy, —S-(1-4C)alkyl, —S(O)-(1-4C)alkyl, —S(O)₂-(1-4C)alkyl,—C(O)O(1-4C)alkyl, carboxy, cyano, hydroxy, nitro, trifluoromethyl andtrifluoromethoxy;

wherein q is an integer from 2 to 6; r is an integer from 1 to 5; and sis an integer from 1 to 5; provided that q+r+s is an integer from 4 to8; and wherein the phen-1,4-ylene group is optionally substituted withfrom 1 to 4 substituents independently selected from halo, (1-4C)alkyl,(1-4C)alkoxy, —S-(1-4C)alkyl, —S(O)-(1-4C)alkyl, —S(O)₂-(1-4C)alkyl,—C(O)O(1-4C)alkyl, carboxy, cyano, hydroxy, nitro, trifluoromethyl andtrifluoromethoxy;

wherein t is an integer from 2 to 10; and u is an integer from 2 to 10;provided that t+u is an integer from 4 to 12;

wherein v is an integer from 2 to 7; and w is an integer from 1 to 6;provided that v+w is an integer from 3 to 8; and wherein thephen-1,4-ylene group is optionally substituted with from 1 to 4substituents independently selected from halo, (1-4C)alkyl,(1-4C)alkoxy, —S-(1-4C)alkyl, —S(O)-(1-4C)alkyl, —S(O)₂-(1-4C)alkyl,—C(O)O(1-4C)alkyl, carboxy, cyano, hydroxy, nitro, trifluoromethyl andtrifluoromethoxy; and

wherein x is an integer from 2 to 6; y is an integer from 1 to 5; and zis an integer from 1 to 5; provided that x+y+z is an integer from 4 to8; and wherein the phen-1,4-ylene group is optionally substituted withfrom 1 to 4 substituents independently selected from halo, (1-4C)alkyl,(1-4C)alkoxy, —S-(1-4C)alkyl, —S(O)-(1-4C)alkyl, —S(O)₂-(1-4C)alkyl,—C(O)O(1-4C)alkyl, carboxy, cyano, hydroxy, nitro, trifluoromethyl andtrifluoromethoxy.
 10. The compound of claim 1, wherein R⁴ is selectedfrom: —(CH₂)₇—; —(CH₂)₈—; —(CH₂)₉—; —(CH₂)₁₀—; —(CH₂)₁₁—;—(CH₂)₂C(O)NH(CH₂)₅—; —(CH₂)₂N(CH₃)C(O)(CH₂)₅—;—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂—; —(CH₂)₂NHC(O)(phen-1,4-ylene)CH₂—;—(CH₂)₂NHC(O)NH(CH₂)₅—; —(CH₂)₃NHC(O)NH(CH₂)₅—;—(CH₂)₂C(O)NHCH₂(cyclohex-1,3-ylene)CH₂—;—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)-;—(CH₂)₂NHC(O)NH(phen-1,4-ylene)(CH₂)₂—;1-[—(CH₂)₂C(O)]piperidin-4-yl)(CH₂)₂—;—(CH₂)₂NHC(O)(trans-cyclohex-1,4-ylene)CH₂—;—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)-;—(CH₂)₂NH(phen-1,4-ylene)(CH₂)₂—;1-[-(CH₂)₂NHC(O)](piperidin-4-yl)(CH₂)₂—;—CH₂(phen-1,4-ylene)NH(phen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)CH₂—;—(CH₂)₂C(O)NHCH₂(pyrid-2,6-ylene)CH₂—;—(CH₂)₂C(O)NH(cis-cyclohex-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(trans-cyclohex-1,4-ylene)CH₂—;—(CH₂)₂NHC(O)(cis-cyclopent-1,3-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)(phen-1,3-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)— ((S)-isomer);—(CH₂)₂C(O)NH(phen-1,4-ylene)C*H(CH₃)— ((R)-isomer);2-[(S)-(—CH₂-](pyrrolidin-1-yl)C(O)(CH₂)₄—;2-[(S)-(—CH₂-](pyrrolidin-1-yl)C(O)(phen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(4-chlorophen-1,3-ylene)CH₂—;—CH₂(2-fluorophen-1,3-ylene)CH₂—;—(CH₂)₂C(O)NH(4-methylphen-1,3-ylene)CH₂—;—(CH₂)₂C(O)NH(6-chlorophen-1,3-ylene)CH₂—;—(CH₂)₂C(O)NH(2-chlorophen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(2,6-dichlorophen-1,4-ylene)CH₂—;—(CH₂)₂NHC(O)NHCH₂(phen-1,3-ylene)CH₂—;4-[—CH₂—](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂—;—(CH₂)₂C(O)N(CH₂CH₃)(phen-1,4-ylene)CH₂—;1-[—(CH₂)₂NHC(O)](piperidin-4-yl)-;—(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₂NHC(O)(thien-2,5-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)(3-nitrophen-1,4-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)(trans-cyclohex-1,4-ylene)-;1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)-;5-[—(CH₂)₂NHC(O)](pyrid-2-yl)CH₂—; —(CH₂)₂(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₃(thien-2,5-ylene)(CH₂)₃—;—(CH₂)₂(phen-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂—;—CH₂(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂—;1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₂—;1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)CH₂—;—(CH₂)₂C(O)NH(3-chlorophen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(2-(CF₃O—)phen-1,4-ylene)CH₂—;—(CH₂)₃(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂—; —(CH₂)₂S(O)₂NH(CH₂)₅—;—CH₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₂C(O)NH(2-iodophen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(2-chloro-5-methoxyphen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(2-chloro-6-methylphen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(CH₂)₅—; —(CH₂)₂N(CH₃)S(O)₂(phen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(2-bromophen-1,4-ylene)CH₂—;—(CH₂)₃(phen-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₃(phen-1,2-ylene)NH(phen-1,4-ylene)(CH₂)₂—;1-[—CH₂(2-fluorophen-1,3-ylene)CH₂](piperidin-4-yl)(CH₂)₃—;—(CH₂)₂C(O)NH(2-methoxyphen-1,4-ylene)CH₂—;—(CH₂)₅NH(phen-1,4-ylene)(CH₂)₂—;4-[—(CH₂)₂-](piperidin-1-yl)(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₂C(O)NH(phen-1,4-ylene)CH(CH₃)CH₂—;—(CH₂)₂-(trans-cyclohex-1,4-ylene)NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₂C(O)NH(2-fluorophen-1,4-ylene)CH₂—;—(CH₂)₂(phen-1,3-ylene)NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₂C(O)NH(2,5-difluorophen-1,4-ylene)CH₂—;—(CH₂)₂NHC(O)(phen-1,4-ylene)(CH₂)₂—;1-[—CH₂(Pyrid-2,6-ylene)CH₂](piperidin-4-yl)CH₂—;—(CH₂)₃NH(phen-1,4-ylene)(CH₂)₂—; —(CH₂)₂NH(naphth-1,4-ylene)(CH₂)₂—;—(CH₂)₃O(phen-1,4-ylene)CH₂—; 1-[—(CH₂)₃](piperidin-4-yl)CH₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)(phen-1,4-ylene)CH₂—;—(CH₂)₃(phen-1,4-ylene)NHC(O)(CH₂)₂—; —(CH₂)₃O(phen-1,4-ylene)(CH₂)₂—;2-[—(CH₂)₂](benzimidazol-5-yl)CH₂—;—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₂—;—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₄—;—(CH₂)₂-(trans-cyclohex-1,4-ylene)NHC(O)(CH₂)₅—;4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₂—;—(CH₂)₂NHC(O)NH(phen-1,4-ylene)CH₂—;—(CH₂)₂N(CH₃)(CH₂)₂(cis-cyclohex-1,4-ylene)-;—(CH₂)₂C(O)NH(2,3,5,6-tetrafluorophen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(2,6-diiodophen-1,4-ylene)CH₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₃—;4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₄—;4-[—(CH₂)₂](piperidin-1-yl)C(O)(CH₂)₅—;—(CH₂)₂C(O)NHCH₂(phen-1,4-ylene)CH₂—;—(CH₂)₂NHC(O)NHCH₂(phen-1,4-ylene)CH₂—;—(CH₂)₂C(O)NH(2-methylphen-1,4-ylene)CH₂—;1-[—(CH₂)₃O(phen-1,4-ylene)(CH₂)₂](piperidin-4-yl)CH₂—;—(CH₂)₂C(O)NHCH₂(phen-1,3-ylene)(CH₂)₂—; —(CH₂)₂O(phen-1,3-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,4-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)CH₂O(phen-1,3-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)(fur-2,5-ylene)CH₂—;—(CH₂)₂N(CH₃)C(O)(thien-2,5-ylene)CH₂—;—(CH₂)₂O(phen-1,4-ylene)O(CH₂)₂—;—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,4-ylene)CH₂—;—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,2-ylene)CH₂—;—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,3-ylene)CH₂—;—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)CH₂O(phen-1,4-ylene)CH₂—;—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂—;—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(thien-2,5-ylene)CH₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,2-ylene)CH₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,3-ylene)CH₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)CH₂O(phen-1,4-ylene)CH₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)(fur-2,5-ylene)CH₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)(thien-2,5-ylene)CH₂—;—(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,3-ylene)CH₂—;—(CH₂)₂(phen-1,4-ylene)NHC(O)(phen-1,4-ylene)CH₂—;—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,2-ylene)CH₂—;—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,3-ylene)CH₂—;—(CH₂)₂(phen-1,4-ylene)NHC(O)CH₂O(phen-1,4-ylene)CH₂—;—(CH₂)₂(phen-1,4-ylene)NHC(O)(fur-2,5-ylene)CH₂—;—(CH₂)₂(phen-1,4-ylene)NHC(O)(thien-2,5-ylene)CH₂—;—(CH₂)₂(trans-cyclohex-1,4-ylene)NHC(O)(phen-1,3-ylene)CH₂—;—(CH₂)₃O(phen-1,3-ylene)CH₂—; —CH₂CH(OH)CH₂NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₄NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₂C(O)NH(phen-1,4-ylene)CH₂NHC(O)CH₂—;—(CH₂)₂C(O)NH(phen-1,4-ylene)(CH₂)₂NHC(O)CH₂—;—(CH₂)₂C(O)NHCH₂(trans-cyclohex-1,4-ylene)CH₂—; —(CH₂)₂NHC(O)(CH₂)₅—;—(CH₂)₂O(phen-1,3-ylene)O(CH₂)₂—; —(CH₂)₂O(phen-1,2-ylene)O(CH₂)₂—;—CH₂(phen-1,2-ylene)O(phen-1,2-ylene)CH₂—; —(CH₂)₂C(O)NH(CH₂)₆—;—(CH₂)₃(phen-1,4-ylene)(CH₂)₃—; —(CH₂)₃(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₄(phen-1,4-ylene)(CH₂)₂—; —(CH₂)₃(furan-2,5-ylene)(CH₂)₃—;—(CH₂)₂N(CH₃)C(O)NH(phen-1,4-ylene)(CH₂)₂—;4-[—(CH₂)₂](piperidin-1-yl)C(O)NH(phen-1,4-ylene)(CH₂)₂—;—(CH₂)₃(phen-1,3-ylene)(CH₂)₃—;—(CH₂)₃(tetrahydrofuran-2,5-ylene)(CH₂)₃—; and—(CH₂)₂O(phen-1,4-ylene)C(O)(CH₂)₂—.
 11. A pharmaceutical compositioncomprising a pharmaceutically acceptable carrier and a therapeuticallyeffective amount of a compound of claim 1.